Appl Environ Microbiol, 1997 Mar, 63(3), 1131 - 8 Characterization of the genes encoding the three-component membrane-bound alcohol dehydrogenase from Gluconobacter suboxydans and their expression in Acetobacter pasteurianus; Kondo K et al.; The three-component membrane-bound alcohol dehydrogenase (ADH) of Gluconobacter suboxydans IFO12528 was purified, and the NH2-terminal amino acid sequence of each subunit was determined . On the basis of the amino acid sequences, the genes adhA, encoding the 72-kDa dehydrogenase, adhB, encoding the 44-kDa cytochrome c-553 (a CO-binding cytochrome c), and adhS, encoding a 15-kDa protein, were cloned and the amino acid sequences of their products were deduced from the nucleotide sequences . The dehydrogenase and cytochrome genes were clustered with the same transcription polarity, as is the case in species of Acetobacter, another genus of acetic acid bacteria . These AdhA and AdhB subunits showed similarity in amino acid sequence to those from Acetobacter spp., whereas AdhS showed no similarity to the corresponding subunit of the ADH complex of Acetobacter pasteurianus . Consistent with this, adhS of G . suboxydans could not complement a defect in the corresponding subunit of A . pasteurianus . When the adhA-adhB gene cluster of G . suboxydans was expressed in an ADH-deficient mutant of A . pasteurianus, the transformant showed distinct ADH activity . The ADH complex was purified to near homogeneity and consisted of two subunits, the dehydrogenase and the cytochrome c subunits derived from G . suboxydans, without any other subunit . These data suggested that AdhS, the smallest subunit of ADH, from G . suboxydans is not essential for ADH activity in A . pasteurianus, in contrast to the essential role of A . pasteurianus AdhS, which is required for correct assembly of the dehydrogenase and cytochrome c subunits on the membrane. J Gen Virol, 1997 Mar, 78 ( Pt 3), 535 - 42 The beet yellows closterovirus p65 homologue of HSP70 chaperones has ATPase activity associated with its conserved N-terminal domain but does not interact with unfolded protein chains; Agranovsky AA et al.; The positive-strand RNA genome of beet yellows closterovirus (BYV) encodes a 65 kDa protein (p65) related to the HSP70 family of cell chaperones . The full-sized BYV p65, and N- and C-terminal fragments, with (His)6 tails, were overexpressed in bacteria and purified by metal-chelate chromatography . Using a polyclonal antiserum raised against the C-terminal fragment of p65, evidence was obtained for expression of the viral protein in planta . Purified recombinant p65 and its N-terminal 40 kDa fragment exhibited Mg2+-dependent ATPase activity in vitro . However, unlike its cellular HSP70 homologues, p65 was unable to bind to denatured protein and its ATPase activity was not stimulated by synthetic peptides which are known to stimulate HSP70 ATPases . Hence, the BYV p65, although being a chaperone-type ATPase, may have a distinct substrate specificity and function in BYV-infected cells. J Bacteriol, 1997 Mar, 179(5), 1628 - 35 Relationship of Treponema denticola periplasmic flagella to irregular cell morphology; Ruby JD et al.; Treponema denticola is an anaerobic, motile, oral spirochete associated with periodontal disease . We found that the periplasmic flagella (PFs), which are located between the outer membrane sheath and cell cylinder, influence its morphology in a unique manner . In addition, the protein composition of the PFs was found to be quite complex and similar to those of other spirochetes . Dark-field microscopy revealed that most wild-type cells had an irregular twisted morphology, with both planar and helical regions, and a minority of cells had a regular right-handed helical shape . High-voltage electron microscopy indicated that the PFs, especially in those regions of the cell which were planar, wrapped around the cell body axis in a right-handed sense . In those regions of the cell which were helical or irregular, the PFs tended to lie along the cell axis . The PFs caused the cell to form the irregular shape, as two nonmotile, PF-deficient mutants (JR1 and HL51) were no longer irregular but were right-handed helices . JR1 was isolated as a spontaneously occurring nonmotile mutant, and HL51 was isolated as a site-directed mutant in the flagellar hook gene flgE . Consistent with these results is the finding that wild-type cells with their outer membrane sheath removed were also right-handed helices similar in shape to JR1 and HL51 . Purified PFs were analyzed by two-dimensional gel electrophoresis, and several protein species were identified . Western blot analysis using antisera to Treponema pallidum PF proteins along with N-terminal amino acid sequence analysis indicated T . denticola PFs are composed of one class A sheath protein of 38 kDa (FlaA) and three class B proteins of 35 kDa (FlaB1 and FlaB2) and one of 34 kDa (FlaB3) . The N-terminal amino acid sequences of the FlaA and FlaB proteins of T . denticola were most similar to those of T . pallidum and Treponema phagedenis . Because these proteins were present in markedly reduced amounts or were absent in HL51, PF synthesis is likely to be regulated in a hierarchy similar to that found for flagellar . synthesis in other bacteria. J Clin Microbiol, 1997 Mar, 35(3), 691 - 6 Aseptic meningitis and encephalitis: the role of PCR in the diagnostic laboratory; Read SJ et al.; In this study we have devised a simple and robust PCR strategy to detect a wide range of viruses, bacteria, and parasites, all of which are capable of causing aseptic meningitis and encephalitis . The techniques developed have been used in a routine diagnostic virology laboratory to test prospectively 2,233 cerebrospinal fluid specimens . A virus was detected in 147 specimens of cerebrospinal fluid from 143 patients . Four sets of primers were sufficient to detect the virus in 135 (94%) of the PCR-positive patients . We conclude that with appropriate primers, PCR can be systematically and economically applied to test for a range of organisms in a routine diagnostic laboratory . In our opinion, PCR will soon become the "gold standard" test for viral infections of the central nervous system. Cytometry, 1997 Mar 1, 27(3), 269 - 74 Flow cytometric determination of endocytosis of viable labelled Legionella pneumophila by Acanthamoeba palestinensis; Harf C et al.; Endocytosis of fluorescently labelled cells of Legionella pneumophila (L . pneumophila) by free-living Acanthamoeba palestinensis (A . palestinensis) has been studied using flow cytometry . L . pneumophila cells were labelled with CM-DiI, a lipophilic fluorescent probe under conditions that did not modify viability . Coculturing the bacteria with amoebae was accompanied by rapid endocytosis; after 5 min, 90% of the amoebae had internalized bacteria . This percentage remained unchanged during further coculture, but the number of bacteria ingested per amoeba increased . Moreover, the number of ingested bacteria was found to be dependent on the size of the amoeba . The validity of the internalization analyzed by flow cytometry was confirmed by observation using epifluorescence and phase contrast microscopy . CM-DiI labelling associated with flow cytometry provides a very valuable technique for the determination of bacteria endocytosis by free-living amoeba. Infect Immun, 1997 Mar, 65(3), 1088 - 94 The Chlamydia trachomatis parasitophorous vacuolar membrane is not passively permeable to low-molecular-weight compounds; Heinzen RA et al.; Chlamydia trachomatis is an obligately intracellular bacterial parasite of eucaryotic cells that undergoes a biphasic life cycle within a parasitophorous vacuole (PV) called an inclusion . The parasitophorous vacuolar membrane (PVM) constitutes a barrier between the replicating bacteria and the nutrient-rich environment of the host cytoplasm . To determine whether the chlamydial PVM contains pores that allow passive diffusion of metabolites between the host cytoplasm and the PV, fluorescent tracer molecules were introduced directly into the cytoplasm of infected cells by transfection or microinjection . Fluorescence microscopy and laser scanning confocal microscopy were subsequently employed to determine whether equilibration of the fluorescent tracers between the cytoplasm and the PV occurred . No movement of tracer molecules as small as 520 Da from the cytoplasm to the PV was observed . These data suggest that the chlamydial PV is not passively permeable to small molecules through open channels in the PVM. J Immunol, 1997 Mar 1, 158(5), 2218 - 27 A three-domain T cell receptor is biologically active and specifically stains cell surface MHC/peptide complexes; Plaksin D et al.; We have expressed in bacteria a single-chain T cell receptor (scTCR) with specificity for an HIV gp120-derived peptide bound to the murine MHC-I molecule, H-2Dd . This scTCR consists of V alpha covalently linked to the VbetaCbeta domains that was solubilized, refolded, and purified in high yield . Specific binding of the scTCR to MHC/peptide complexes was demonstrated by surface plasmon resonance, with a Kd of 2 to 8 x 10(-6) M . This scTCR specifically inhibited T cell activation, and stained cell surface MHC/peptide complexes as measured by cytofluorimetry . The preservation of binding specificity by such a three-domain scTCR suggests that this structure is sufficient for specific MHC/peptide recognition and that this strategy will be of general use as applied to other TCR. Sci Total Environ, 1997 Feb 24, 194-195, 247 - 62 Biology of the Humber rivers; Whitton BA et al.; An overview of the literature is presented on the biology of the rivers entering the Humber, eastern England, together with some of their tributaries . Particular emphasis is given to dynamic aspects, including transport and movement within rivers, movement between rivers, processes within rivers and long-term changes. Presse Med, 1997 Feb 22, 26(5), 248 - 54 {Search for a HIV vaccine}; Sicard D et al.; A DIFFICULT SITUATION FOR A VACCINE: The human immunodeficiency virus has an exceptional capacity to mutate and macrophage reservoirs where it is harbored after penetration are highly inaccessible to antibodies . Use of a live vaccine would increase the risk of recurrent virulence and integration into the genome . UNKNOWN IMMUNOLOGY: Induction of cytotoxic cells appears to be essential, but investigations into this type of response are not well standardized and not easily quantifiable . Neutralizing antibodies would have a protective effect, but facilitating antibodies would have the opposite effect . SEVERAL POSSIBILITIES UNDER STUDY: These vaccine use live attenuated viruses with a more or less deleted genome, recombinant live viruses constructed from other viruses or bacteria, pseudo-particles, or recombinant proteins as well as vaccines synthetized from peptides or lipopeptides . An association of different vaccines would appear to be the best solution as live vectors usually induce cytotoxic cells while proteins or peptides induce production of neutralizing antibodies . RESPONSE TO VACCINES IS USUALLY WEAK: Both in animal models and in human volunteers, immune responses obtained to date have been quite variable and of short duration . Canarypox type recombinant vaccines followed by recombinant protein vaccines appear to give the best results at the present time . SEVERAL PROBLEMS: Anti-HIV vaccination protocols raise major ethical problems for the participating volunteers (ELISA test becomes positive, lack personal protection) and for the population involved in phase II/III trials . In addition, the virus rapidly penetrates via the mucosa without yielding sufficient mucosal immune response. J Theor Biol, 1997 Feb 21, 184(4), 451 - 69 A study of oligonucleotide occurrence distributions in DNA coding segments; Castrignano T et al.; In this paper we present a general strategy designed to study the occurrence frequency distributions of oligonucleotides in DNA coding segments and to deal with the problem of detecting possible patterns of genomic compositional inhomogeneities and disuniformities . Identifying specific tendencies or peculiar deviations in the distributions of the effective occurrence frequencies of oligonucleotides, with respect to what can be a priori expected, is of the greatest importance in biology . Differences between expected and actual distributions may in fact suggest or confirm the existence of specific biological mechanisms related to them . Similarly, a marked deviation in the occurrence frequency of an oligonucleotide may suggest that it belongs to the class of so-called "DNA signal (target) sequences" . The approach we have elaborated is innovative in various aspects . Firstly, the analysis of the genomic data is carried out in the light of the observation that the distribution of the four nucleotides along the coding regions of the genoma is biased by the existence of a well-defined "reading frame" . Secondly, the "experimental" numbers found by counting the occurrences of the various oligonucleotide sequences are appropriately corrected for the many kinds of mistakes and redundancies present in the available genetic Data Bases . A methodologically significant further improvement of our approach over the existing searching strategies is represented by the fact that, in order to decide whether or not the (corrected) "experimental" value of the occurrence frequency of a given oligonucleotide is within statistical expectations, a measure of the strength of the selective pressure, having acted on it in the course of the evolution, is assigned to the sequence, in a way that takes into account both the value of the "experimental" occurrence frequency of the sequence and the magnitude of the probability that this number might be the result of statistical fluctuations . If the strength of the selective pressure evaluated in this fashion turns out to be sufficiently large, the corresponding sequence will be considered to have an occurrence frequency beyond expectations and, hence, to be statistically and biologically interesting. J Mol Biol, 1997 Feb 21, 266(2), 306 - 16 Mutant RecA proteins which form hexamer-sized oligomers; Logan KM et al.; We have analyzed the oligomeric properties of a number of mutant RecA proteins containing single amino acid substitutions within one region of the subunit interface . In contrast to wild-type RecA, which forms a heterogeneous population of different-sized oligomers, we find that many of these mutant proteins exist in a more homogeneous oligomeric form, which approximates to the size of a RecA hexamer . Some of these mutants have a significant level of activity in vivo for recombinational DNA repair and thus represent the first mutant RecA proteins identified which retain activity yet can exist in a discrete oligomeric state as free protein. Circulation, 1997 Feb 18, 95(4), 814 - 7 Left ventricular assist device infection is associated with increased mortality but is not a contraindication to transplantation; Herrmann M et al.; BACKGROUND: Left ventricular assist devices (LVADs) are increasingly used as a bridge to transplantation . Infection is a frequent and major complication associated with the use of these devices, however, the correlation of infection and outcome has not yet been evaluated in a prospective fashion . METHODS AND RESULTS: Twenty-five patients (24 male, 1 female) with end-stage cardiac failure and resulting organ dysfunction were included . Patients were bridged with the Novacor N100 portable LVAD (median duration of support, 55 days) and were evaluated prospectively by device surface cultures on explantation, molecular typing of isolates, and correlation of infection with survival to transplant . Twelve (48%) of 25 patients had LVAD infection as defined by recovery of multiple isolates of identical genotype from the device surface . Whereas only 5 (42%) of 12 patients with LVAD infection survived until transplantation, 11 (85%) of 13 patients without infection were successfully transplanted (P < .05) . Death of the 7 patients with proven LVAD infection was associated with multiple organ failure or other signs of acute infection . CONCLUSIONS: LVAD infection is associated with a significantly decreased survival probability . It does not preclude successful bridging but rather may pose an indication for urgent transplantation. Biochem J, 1997 Feb 15, 322 ( Pt 1), 207 - 11 Difference in hepatic metallothionein content in Antarctic red-blooded and haemoglobinless fish: undetectable metallothionein levels in haemoglobinless fish is accompanied by accumulation of untranslated metallothionein mRNA; Scudiero R et al.; Icefish (family Channichthyidae, suborder Nothothenioidei) are a group of Antarctic fish that have evolved unique phenotypes in order to adapt to the environment in which they live . Besides the lack of haemoglobin and the drastic reduction in the number of erythrocyte-like cells, another striking feature of the icefish is that their liver is devoid of metallothionein . These cysteine-rich heavy-metal-binding proteins are usually present in large amounts in a large variety of organisms, from bacteria to mammals . Despite the failure to detect appreciable levels of metallothionein in icefish liver, a cDNA encoding metallothionein was produced from total RNA by reverse transcriptase PCR . The icefish metallothionein showed high percentage identity with metallothionein from Trematomus bernachii, a red-blooded Antarctic fish in which a normal content of hepatic metallothionein was found . Steady-state mRNA levels were assessed in fish liver by high-stringency hybridization of the metallothionein probe with total RNA . The results showed that icefish livers retain large amounts of untranslated metallothionein mRNA . The stability of the icefish transcript might be correlated with the lack of specific motifs in the untranslated 3' ends of mRNA. Anal Biochem, 1997 Feb 15, 245(2), 115 - 22 Recombinant strategies for rapid purification of catalytic subunits of cAMP-dependent protein kinase; Hemmer W et al.; Knowledge of the crystal structure of the catalytic subunit (C) of cAMP-dependent protein kinase provided for the first time a molecular basis for probing function by site-directed mutagenesis . The purification of mutant C-subunits, however, presented new and unanticipated challenges due to instability, insolubility, and underphosphorylation of the altered proteins . To overcome these barriers, a rapid and efficient method for purifying recombinantly expressed C-subunits was developed . Purification to near homogeneity is achieved in less than 5 h . The procedure is based on colysis of bacteria that overexpress the C-subunit with bacteria that overexpress a poly-His-tagged mutant of the type II regulatory subunit H6RII (R213K) . This mutant R-subunit with an altered cAMP binding site A forms holoenzyme rapidly in bacterial extracts, and the Ka (cAMP) for the resulting holoenzyme, 27-37 microM, is nearly 50-fold increased compared to holoenzyme formed with wild-type RII . Thus, after batchwise immobilizing the holoenzyme on Ni(2+)-resin, the free C-subunit can be directly eluted batchwise with high concentrations of cAMP . The method is described for the purification of wild-type C, with yields of approximately 5 mg/liter . In addition, a mutant subunit, C{G52S}, which is defective in ATP binding and could not be isolated using previously described methods, was purified with equal efficiency. Transplantation, 1997 Feb 15, 63(3), 407 - 14 Effects of hypoxemia on early postoperative course of liver transplantation in pediatric patients with intrapulmonary shunting; Uemoto S et al.; Nine pediatric patients (mean age, 10 years) with biliary atresia, who had hypoxemia related to intrapulmonary shunting, underwent living related liver transplantation . The effects of hypoxemia during the early postoperative period after liver transplantation on cardiopulmonary and renal function, as well as on transplanted liver, were analyzed . Based on the degree of shunt ratio calculated by technetium-99m macroaggregated albumin scintigraphy, the nine patients were included in the moderate group (shunt ratio under 40%, n=4) or the severe group (shunt ratio over 40%, n=5) . Partial pressure of arterial oxygen was maintained at normal range in the moderate group, while that in the severe group persistently had very low values (<50 mmHg), in spite of a high degree of oxygen supply . However, all patients in the severe group maintained stable cardiopulmonary vital signs, including systemic blood pressure, heart rate, respiratory rate, and cardiac index . They also demonstrated stable renal function . None of the patients died of cardiopulmonary or renal insufficiency after transplantation, but three patients died of portal vein thrombosis, sepsis, and intracranial hemorrhage (one each) . The minimal adverse effect of hypoxemia on the transplanted liver was confirmed by a rapid increase of arterial ketone body ratio, low peak values (under 200 IU/L) of aspartate aminotransferase, and a steady decrease of serum total bilirubin . Four patients encountered surgical complications, including two bile leaks from the cut liver surface, two leaks from bilioenteric anastomosis, and one intestinal perforation . Six patients suffered from bacterial infections, including four wound infections, three right subphrenic abscesses, one cholangitis, and two systemic sepses . All patients in the moderate group recovered from hypoxemia, but four of five patients in the severe group have not recovered during the follow-up period between 4 and 9 months . It was concluded that the adverse effects of hypoxemia on cardiopulmonary and renal function and transplanted liver were minimal, so that patients with severe hypoxemia could tolerate the stress of liver transplantation without special management . However, the high incidence of surgical complication and infection suggested the adverse effects of hypoxemia on wound healing and resistance to bacteria infection. Structure, 1997 Feb 15, 5(2), 153 - 7 A good turn for DNA: the structure of integration host factor bound to DNA; Ellenberger T et al.; The crystal structure of integration host factor (IHF) complexed with DNA shows how a small heterodimeric protein can induce a big bend in DNA . IHF exerts leverage in the minor groove and wraps DNA around the body of the protein, providing another example of sequence-specific recognition of the minor groove. J Biol Chem, 1997 Feb 14, 272(7), 4058 - 64 Molecular mechanism of polyamine stimulation of the synthesis of oligopeptide-binding protein; Igarashi K et al.; Polyamine stimulation of the synthesis of oligopeptide-binding protein (OppA) was shown to occur mainly at the level of translation by measuring OppA synthesis and its mRNA level . Several artificial oppA genes were constructed by site-directed mutagenesis . These synthesize different kinds of OppA mRNAs: mRNAs differing in the size of 5'-untranslated region; mRNAs having the Shine-Dalgarno (SD) sequence in a different position; mRNAs having different secondary structure in the region of the SD sequence; and fusion mRNAs consisting of the 5'-untranslated region of OppA mRNA and the open reading frame of beta-galactosidase . By measuring the synthesis of OppA or beta-galactosidase from these mRNAs, we found that the 171-nucleotide 5'-untranslated region and 145 nucleotides of the ORF of OppA mRNA are involved in the polyamine stimulation of OppA synthesis . When the secondary structure of the above region of OppA mRNA was analyzed by optimal computer folding, it was shown that the degree of polyamine stimulation of OppA protein synthesis was dependent on the structure of the SD sequence in addition to its position . Loose base pairing of the SD sequence with other regions of the mRNA caused strong polyamine stimulation, while intense base pairing of the SD sequence with other regions of the mRNA resulted in insignificant or weak polyamine stimulation. Proc Natl Acad Sci U S A, 1997 Feb 4, 94(3), 802 - 7 Binding of SecB to ribosome-bound polypeptides has the same characteristics as binding to full-length, denatured proteins; Randall LL et al.; The interaction of the chaperone SecB with ribosome-bound polypeptides that are in the process of elongation has been studied using an in vitro protein synthesis system . The binding is characterized by the same properties as those demonstrated for the binding of SecB to full-length proteins that are in nonnative conformation: it is readily reversible and has no specificity for the leader peptide . In addition, it is shown that the growing polypeptide chains must achieve a critical length to bind tightly enough to allow their isolation in complex with SecB . This explains the longstanding observation that, even when export is cotranslational, it begins late in synthesis . Furthermore, the required length is approximately the same as the length that defines the binding frame within denatured, full-length proteins bound to SecB. Am J Perinatol, 1997 Feb, 14(2), 103 - 6 A pilot study identifying type V collagenolytic activity in human amniotic fluid; Polzin WJ et al.; Amniorrhexis complicates pregnancies if it occurs in a preterm pregnancy or remote from the onset of labor in a term pregnancy . There are different collagen types (I-V) that create the extracellular matrix of the amnion . Collagenases specific to these collagen types, with the exception of type V collagen, are found in human amniotic fluid, fibroblasts, polymorphonuclear leukocytes and bacteria . Type V collagen is a major component of the amniotic basement membrane and is responsible for maintaining a barrier to bacteria and to the loss of amniotic fluid . We sought to find evidence of type V collagenolytic activity in human amniotic fluid obtained from pregnancies in different clinical states. Endod Dent Traumatol, 1997 Feb, 13(1), 31 - 5 Experimental apexigenesis in baboons; Das S et al.; Apexification with calcium hydroxide is a routine procedure . However, some clinical reports suggest that root completion can occur by controlling the infection without use of a catalyst . The present study investigated the use of tetracycline treatment (in root canals) on root growth in immature teeth, rendered non-vital experimentally . Incisors in 3 young baboons were exposed and canals were left open . After 2 months all canals were cleaned and treated with either tetracycline or formocresol . Some canals in each group were filed . Animals were sacrificed after 6 months . Bacterial evaluations were done before placing medications, one week later and six months after that . The number of bacteria were reduced in all treatment groups . Root growth almost near completion was observed in more teeth treated with tetracycline than in the formocresol group. J Infect Dis, 1997 Feb, 175(2), 474 - 7 Interleukin-8 and chemotactic activity of middle ear effusions; Storgaard M et al.; The importance of interleukin (IL)-8 in the chemotactic activity of middle ear effusions (MEEs) was evaluated . There was a significantly higher IL-8 concentration in MEEs of children with acute otitis media (AOM) (n = 17; 136 ng/mL) than in children with otitis media with effusion (OME) (n = 28; 65 ng/mL) . The IL-8 concentration in MEEs with bacteria (149 ng/mL) was significantly higher than in MEEs without bacteria (66 ng/mL) . MEEs from children with AOM and OME had equally higher chemotactic activity than the diluent alone (23.3% and 24.8% vs . 9.2%) . The chemotactic activity was not altered by the presence of bacteria nor did it correlate with IL-8 concentration . Fractionation of MEEs by gel chromatography demonstrated that the main chemotactic activity could clearly be separated from the IL-8 activity, thus excluding IL-8 as a main chemotactic component in MEEs. Eur J Cancer, 1997 Feb, 33(2), 209 - 13 Bile duct stents: is there an increased rate of complications in patients receiving chemotherapy? Lofts FJ, Evans TR, Mansi JL, Glees JP, Knight MJ. The aim of this study was to determine whether palliative chemotherapy accelerates the rate of biliary stent occlusion, in patients with a malignant biliary obstruction . Such treatment can induce neutropenia and increase the risk of bacterial sepsis . Overgrowth of bacteria within the bile of patients receiving chemotherapy could accelerate the rate of stent occlusion . Retrospective analysis of treatment records for 80 consecutive patients with a diagnosis of adenocarcinoma arising from the pancreas, bile ducts or gall bladder was conducted . Two groups were identified, those with a biliary stent in situ (primary stent group: 47/80; 59%) at the time of referral and those without (no stent group: 33/80; 41%) . The majority of patients went on to receive chemotherapy, 64% and 70% in the primary stent group and no stent group, respectively . The rate of febrile neutropenia was similar in the two groups (5% versus 7% of all chemotherapy cycles in the primary stent group and no stent group, respectively) . The rate of stent occlusion was not significantly different between those exposed to chemotherapy (37%; 95% CI 20-54%) and those unexposed (39%; 95% CI 19-59%) . Similarly, the mean duration of patency was not shortened by chemotherapy (105 days in the chemotherapy group versus 119 days in the non-chemotherapy group; P = 0.97, Mann-Whitney U-test) . We conclude that there is no evidence of increased rate of bile duct-related complications in patients receiving chemotherapy . In particular, we find no indication for the use of prophylactic antibiotics. J Comp Pathol, 1997 Feb, 116(2), 203 - 16 An immunohistochemical study of the lesions of demodicosis in the dog; Day MJ; Histopathological and immunohistochemical examination of skin biopsies from 32 dogs with demodicosis is reported . There was no association between the different clinical presentations of the disease and the histopathological character of the biopsies, which included absence of inflammation (n = 2), dominant perifolliculitis (n = 11), interface mural folliculitis (n = 7), mural folliculitis (n = 1), furunculosis (n = 10) and nodular dermatitis (n = 1) . In eight of 32 biopsies colonies of coccoid bacteria or Malassezia pachydermatis-like yeasts were observed . IgG-bearing plasma cells were found in similar numbers in the inflammatory infiltrates of all types of histological lesion, and were invariably more numerous than IgM or IgA plasma cells . The IgG plasma cells were largely IgG4 in lesions of perifolliculitis, but consisted of a mixture of IgG2 and IgG4 where folliculitis or furunculosis was present . CD3+T lymphocytes were prominent within the interface infiltrates of follicular epithelium and also within the lesions of furunculosis . Dermal inflammatory cells and epidermal Langerhans cells expressing MHC Class II were observed in similar number in all types of lesion . The study demonstrated an active local cutaneous immune response in canine demodicosis, which increased as the dermal pathology progressed from perifolliculitis to furunculosis. Acta Paediatr Jpn, 1997 Feb, 39(1), 105 - 13 Trends in development of anti-infective agents in Japan; Yagisawa M et al.; Newly developed anti-infective agents have been continuously supplied in the clinics of Japan over the past 50 years, and were beneficial in saving patients from life-threatening infections . However, the emergence of resistant bacteria and uncommon pathogens has caused complications in chemotherapy . Further efforts have been made to develop newer and more effective agents to minimize such complications . The newest and the most effective agents are available even at primary-care clinics, as the health insurance system allows the use of any agents approved by the government . Despite potential risks of the emergence and spread of resistant bacteria and the occurrence of adverse drug reactions, current usage of such effective agents shows more successful results, particularly in prognosis, than that of less effective ones . Because of the extreme changes in infectious diseases over the last 10-15 years, guidelines for the evaluation of anti-infective agents have been proposed . It is essential to harmonize with the USA and the European guidelines for mutual acceptance of clinical data . However, it is true that practices of anti-infective therapy in Japan, in the USA and in Europe are different both in dose regimen and assessments of efficacy and safety . As the leading country in the development of newer anti-infective agents, Japan proposes the most sophisticated way to evaluate those agents to update knowledge in medical science. Nutrition, 1997 Feb, 13(2), 128 - 32 Effect of total parenteral nutrition with different lipid emulsions of human monocyte and neutrophil functions; Waitzberg DL et al.; Parenteral nutrition (TPN) with lipid emulsions is claimed to be associated with impaired monocyte (M) and neutrophil (N) functions . Long-chain triglycerides (LCT) and a mixture containing 50% medium-chain triglycerides (MCT) and 50% LCT, currently used in nutritional therapy with TPN, were evaluated for their ex vivo effects on human N and M chemotaxis, phagocytosis, bacterial killing, and oxidative metabolism by nitroblue tetrazolium reduction test . Cell functions were examined in a randomized, crossover, blind trial in 10 malnourished patients with gastric cancer . Prior to the operation (2 wk), central TPN (40 kcal/kg) with 25% of caloric energy provided as LCT or MCT/LCT emulsion was infused over 48 h . After the crossover period fat-free TPN was given over 48 h . Function tests were done for N and M before and after each lipid emulsion infusion . Every cell function test performed for each patient was controlled by another test done in healthy adult volunteers and the results were compared with the normal range of values previously established for a healthy adult population . All the patients completed the studies without complications . Crossover validity was statistically established . Bacterial killing was the only function reduced in neutrophils after LCT emulsion (% killed bacteria = 79.0 +/- 8.5 versus 67.4 +/- 19.2; P < 0.05), although this function remained within the normal range values in 80% of the patients . In conclusion, the lipid emulsions did not affect any monocyte functions and only moderately decreased neutrophil bacterial killing. J R Army Med Corps, 1997 Feb, 143(1), 31 - 4 Dermatological conditions in winter in primary health care on Operation Resolute (Bosnia); Winfield DA; The distribution of dermatological conditions has been studied in a total of 1822 consultations with British troops in a primary health care setting on Operation Resolute (Bosnia) between 1 January and 4 March 1996 . Approximately one in eight (12%) of the consultations were for skin conditions; eczema was the most common complaint, but, taken as a whole, infections due to virus (excluding warts), fungus and bacteria made up 30% . The overall distribution of diseases was similar to that seen in British general practice. Int J Parasitol, 1997 Feb, 27(2), 173 - 80 Endoparasite control strategies: implications for biodiversity of native fauna; Spratt DM; Efforts to control the spectrum of diseases that affect humans, our crops and our animals pose problems which need to be debated openly . Widespread use of chemicals in such a broad sphere raises important concerns not only about safety for the users, consumers and target species, but especially about the not so obvious effects upon the ecosystems in which they are used . Some undetermined level of biological diversity is necessary to maintain ecological function and resilience . These, in turn, are necessary for generating the biological resources (trees, fish, wildlife, crops) and ecological services (watershed protection, air cleansing, climate stabilisation, erosion control) on which economic activity and human welfare depend . The driving forces behind decline of biodiversity stem entirely from human activities . Underlying causes are those resulting from the cultural and social factors associated with economic activities and lead to direct depletion of species, and degradation or destruction of habitats . The broad spectrum and high efficacy of the macrocyclic lactones against nematode and arthropod parasites of livestock and companion animals are unprecedented . Cattle, horses, sheep, swine, dogs--to varying degrees all are utilised by humans for economic gain . Detrimental impact upon non-target animals is considered acceptable in eradicating parasites because of their economic importance to commercial livestock production . Production will increase when these parasites are eliminated, but we remain oblivious to the long-term consequences of our actions . What are the ecological limits to rural economic activities? Decomposing animal faeces help to maintain our ecosystem by returning valuable nutrients to the soil . Dung fauna-fungi, yeast, bacteria, nematodes, insects and earthworms--play a non-conspicuous but important and varied role in this decomposition process, a role dependent upon many factors, especially environmental ones . Anthelmintics and pesticides are of considerable value in agriculture, but largely at an unevaluated cost to the greater environment . We have insufficient knowledge of the extent to which a spectrum of anthelmintics and pesticides affect ecological function and ecosystem resilience in our commercial plant and animal production systems . It is time we developed a genuine interest in avoiding "the dialogue of the deal" that in the past has minimised interdisciplinary research between environmental ecology and commercial plant and animal production. Hybridoma, 1997 Feb, 16(1), 11 - 6 Combinatorial antibodies against human malignant melanoma; Pereira S et al.; The general responsiveness of human melanoma to immunotherapy has been well established, but active immunotherapy of melanoma has been hampered by insufficient information on the immunogenicity of melanoma antigens in patients . We have attempted to identify melanoma-associated antigens recognized by patients' B cells using an antibody phage display approach . Antibody display on filamentous phages allows direct screening of cDNA libraries for expression of cell-surface-reactive antibodies, without the need for antibody production and purification using bacteria or eukaryotic cell systems . This approach was used to identify melanoma-associated cell-surface antigens recognized by patients' B cells . Antibodies produced by the B cells of a melanoma patient (in remission for > 7 years following periodic vaccination with allogeneic melanoma cell vaccine) were displayed as Fabs on the surfaces of filamentous phages . A library of 10(8) phages was absorbed to normal melanocytes, followed by phage binding to and elution from melanoma cells (human lymphocyte antigen nonmatched and vaccine melanoma cells) . Phages were further selected for reactivities with tunicamycin-treated melanoma cells . These procedures resulted in a > 10(6)-fold enrichment of tumor-specific phages from the original phage library . One phage-Fab bound to melanoma cells, other tumor cells, and a few normal cells in cultured cell lines and in tissue sections. Curr Biol, 1997 Feb 1, 7(2), R108 - 11 Cloning in silico; Cutler S et al.; Cellulose is a major component of plant cell walls, but identification of the enzymes that synthesize it has proven difficult . Now, however, several candidate proteins with sequence homology to bacterial cellulose synthases have been identified by partial sequencing of anonymous cDNA clones from cotton fibers. Gut, 1997 Feb, 40(2), 211 - 4 Chemical synthesis of nitric oxide in the stomach from dietary nitrate in humans; McKnight GM et al.; BACKGROUND/AIMS: It has been suggested that dietary nitrate, after concentration in the saliva and reduction to nitrite by tongue surface bacteria, is chemically reduced to nitric oxide (NO) in the acidic conditions of the stomach . This study aimed to quantify this in humans . METHODS: Ten healthy fasting volunteers were studied twice, after oral administration of 2 mmol of potassium nitrate or potassium chloride . Plasma, salivary and gastric nitrate, salivary and gastric nitrite, and gastric headspace NO concentrations were measured over six hours . RESULTS: On the control day the parameters measured varied little from basal values . Gastric nitrate concentration was 105.3 (13) mumol/l (mean (SEM), plasma nitrate concentration was 17.9 (2.4) mumol/l, salivary nitrate concentration 92.6 (31.6) mumol/l, and nitrite concentration 53.9 (22.8) mumol/l . Gastric nitrite concentrations were minimal (< 1 mumol/l) . Gastric headspace gas NO concentration was 16.4 (5.8) parts per million (ppm) . After nitrate ingestion, gastric nitrate peaked at 20 minutes at 3430 (832) mumol/l, plasma nitrate at 134 (7.2) mumol/l, salivary nitrate at 1516.7 (280.5) mumol/l, and salivary nitrite at 761.5 (187.7) mumol/l after 20-40 minutes . Gastric nitrite concentrations tended to be low, variable, and any rise was non-sustained . Gastric NO concentrations rose considerably from 14.8 (3.1) ppm to 89.4 (28.6) ppm (p < 0.0001) after 60 minutes . All parameters remained increased significantly for the duration of the study . CONCLUSIONS: A very large and sustained increase in chemically derived gastric NO concentrations after an oral nitrate load was shown, which may be important both in host defence against swallowed pathogens and in gastric physiology. Genetics, 1997 Feb, 145(2), 467 - 78 Transposon-disruption of a maize nuclear gene, tha1, encoding a chloroplast SecA homologue: in vivo role of cp-SecA in thylakoid protein targeting; Voelker R et al.; A nuclear mutant of maize, tha1, which exhibited defects in the translocation of proteins across the thylakoid membrane, was described previously . A transposon insertion at the tha1 locus facilitated the cloning of portions of the tha1 gene . Strong sequence similarity with secA genes from bacteria, pea and spinach indicates that tha1 encodes a SecA homologue (cp-SecA) . The tha1-ref allele is either null or nearly so, in that tha1 mRNA is undetectable in mutant leaves and cp-SecA accumulation is reduced > or = 40-fold . These results, in conjunction with the mutant phenotype described previously, demonstrate that cp-SecA functions in vivo to facilitate the translocation of OEC33, PSI-F and plastocyanin but does not function in the translocation of OEC23 and OEC16 . Our results confirm predictions for cp-SecA function made from the results of in vitro experiments and establish several new functions for cp-SecA, including roles in the targeting of a chloroplast-encoded protein, cytochrome f, and in protein targeting in the etioplast, a nonphotosynthetic plastid type . Our finding that the accumulation of properly targeted plastocyanin and cytochrome f in tha1-ref thylakoid membranes is reduced only a few-fold despite the near or complete absence of cp-SecA suggests that cp-SecA facilitates but is not essential in vivo for their translocation across the membrane. Hum Reprod, 1997 Feb, 12(2), 286 - 91 Fertilization with human testicular spermatids: four successful pregnancies; Antinori S et al.; Between July 1995 and May 1996, 36 patients with non-obstructive azoospermia of secretory origin underwent intracytoplasmic injection of spermatids . A previous histological biopsy was performed on all patients: 15 had spermatogenic arrest, a further 13 had Sertoli cell-only syndrome, and the remaining eight had post-cryptorchidism tubal atrophy . The ejaculate was duly examined and a complete absence of spermatozoa and spermatids was confirmed, with only bacteria and debris being found . Testicular sperm extraction (TESE) was then performed . In 19 out of 36 cases round spermatids only were found, while elongated spermatids were found in the remaining 17 . Both round and elongated spermatids were isolated and used for injection . A total of 135 oocytes at metaphase II were recovered from 19 partners and injected with round spermatids, while 123 mature oocytes from 17 partners were injected with elongated spermatids . The number of oocytes fertilized, as judged by the presence of two pronuclei, was 75 (55.5%) and 71 (57.7%) respectively . By 34 h after injection, the number of embryos which had cleaved to the 2-cell stage was 56 (74.6%) with round spermatids and 55 (77.4%) with elongated spermatids . All cleaved embryos were transferred into the uterus of the partners . Clinical pregnancies were established in two cases of round spermatid cycles (10.5%) (both are still ongoing), and three cases of elongated spermatid cycles (17.6%) (two are still ongoing; one was lost after 8 weeks of gestation) . Chromosomal analysis showed that all fetuses had a normal karyotype (three male and one female) with no chromosomal abnormalities. Surg Endosc, 1997 Feb, 11(2), 93 - 4 Thoracoscopic decortication in infants and children; Rothenberg SS et al.; BACKGROUND: The treatment of pediatric empyemas remains controversial . While thoracentesis and tube thoracostomy appear adequate for relatively benign organisms, virulent bacteria cause thick fibrinous pleural peels entrapping the lung . Open thoracotomies have been effectively used for decortication but are painful . METHODS: We report the use of minimally invasive thoracoscopic decortication in 12 patients (mean age 5 years) . All failed conventional management with persistent fever, increasing oxygen requirement, recurrent effusion, and pleural consolidation; 5- and 10-mm trocars were used and complete decortication was accomplished . RESULTS: Ten of 12 patients were afebrile by 72 h and discharged 4-12 days after surgery . Eleven of 12 patients had clear chest x-rays by 1 month . CONCLUSION: Thoracoscopic decortication is a safe and effective means of treating pediatric empyemas. Br J Dermatol, 1997 Feb, 136(2), 260 - 3 Skin infection caused by Mycobacterium avium; Ichiki Y et al.; A patient with skin infection due to Mycobacterium avium is reported . A 9-year-old female had 10 subcutaneous nodules and two ulcers on the abdomen and legs . She had no medical history of systemic disease, skin disease or immunosuppressive therapy . Cultures of a biopsy specimen and of aspirated seropurulent fluid in nodules showed acid-fast bacteria, identified as M . avium by the DNA-DNA hybridization method . We treated her with a combination of surgery and the antibiotics, cycloserine, isoniazid and clarithromycin. Biochem Mol Biol Int, 1997 Feb, 41(2), 243 - 55 The growth kinetics of intracellular bacille Calmette-Guerin (BCG) within human monocyte-derived macrophages of different ethnic populations; Fazal N; In this paper we have examined intracellular mycobacterial growth in human monocyte-derived macrophage cell cultures . Cells from three population groups were selected to compare and contrast the kinetics of intracellular BCG growth both by determining changes in metabolic activity of dividing bacteria by radio-isotope incorporation and growth/survival by estimating colony forming unit by Fazal's Microcolony Counting Method . These data indicate that BCG infects and multiplies intracellularly in human macrophages and may serve as an in vitro model for studying inherent capability of mycobacteria to grow and infect macrophage populations . There is however, no inhibition or reduction of intracellular mycobacterial growth from a Caucasian male and female, and an Afrocarribean female donor macrophages . On the contrary, there is a ten-fold increase in the replication of mycobacteria over a 7-day infection period . Patterns of BCG growth within macrophage cultures from different donors were determined and variation calculated at different days post-infection . Experiment to experiment coefficient of variation of intracellular BCG growth estimates for a single donor is between 4-12% . In contrast, the coefficient of variation of intracellular BCG growth between three different macrophage donors studied is < 4%. Mol Med Today, 1997 Feb, 3(2), 61 - 8 Microsatellite instability in human solid tumors; Lothe RA; A genome-wide instability has been found in almost all analyzed malignant tumors from patients with hereditary non-polyposis colorectal cancer (HNPCC), and in a subgroup of sporadic (non-inherited) cancers of the same type . This mutator phenotype was initially seen as novel alleles at microsatellite loci (a family of repetitive DNA sequences) and was shown to be caused by mutations in the highly conserved mismatch repair genes . Mutations have been found in each of four of these human genes: hMSH2, hMLH1, hPMS1 and hPMS2, in the germline of HNPCC patients and in their tumors, as well as in sporadic tumors . These recent discoveries provide new molecular diagnostic tools for the detection of patients at high risk of developing carcinomas of the large bowel and other HNPCC-related tumors . Ongoing international research is progressively solving many of the unanswered questions at the genotypic and phenotypic levels of this newly identified mechanism in carcinogenesis. J Interferon Cytokine Res, 1997 Feb, 17(2), 69 - 75 Treatment of an infected murine macrophage cell line (J774A.1) with interferon-gamma but not tumor necrosis factor-alpha or live Mycobacterium intracellulare alone modulates the expression of adhesion molecules; Pourshafie MR et al.; In the present study, we evaluated whether the activation of a murine macrophage cell line (J774.1A) by treatment with recombinant murine tumor necrosis factor-alpha (rTNF-alpha) or recombinant murine interferon-gamma (rIFN-gamma) before or simultaneous with infection with Mycobacterium intracellulare would affect their ability to express lymphocyte function-associated antigen-1 (LFA-1) and to restrict growth and kill the ingested M . intracellulare . The data showed that the effect of lipopolysaccharide (LPS) in increasing the level of LFA-1 was the same in the presence or absence of M . intracellulare . The inability of M . intracellulare to affect the level of expression of LFA-1 was irrespective of the M . intracellulare to J774A.1 ratio . A significant increase in the expression of LFA-1 was observed when J774A.1 cells were prestimulated with IFN-gamma 1 day before the addition of the bacteria . The addition of IFN-gamma with M . intracellulare simultaneously, however, did not affect the expression of the adhesion molecules as compared with the IFN-gamma alone . Our results indicated no change in the level of LFA-1 on J774A.1 following exposure with TNF-alpha . We observed that preexposure with 10-10(4) IU/ml of TNF-alpha can significantly decrease the number of ingested M . intracellulare . Simultaneous addition of 10(3) and 10(4) IU/ml of TNF-alpha, however, did not have any mycobactericidal effect . This indicates that the TNF-alpha-induced killing by J774A.1 cells was relatively selective, depending on the concentration and the time of presence of TNF-alpha . The data may suggest that the uptake of M . intracellulare is carried out via other adhesion receptors when M . intracellulare and IFN-alpha are present simultaneously and that in the presence of TNF-alpha other surface receptors are involved in the uptake of M . intracellulare . Flow cytometry analysis of the spleen cells removed at various times from M . intracellulare-infected mice also indicated no change in the level of LFA-1 beta or MAC-1, a finding comparable with that of the J774A.1 cells. Biol Pharm Bull, 1997 Feb, 20(2), 127 - 33 Characterization of lactoferrin-binding proteins of human macrophage membrane: multiple species of lactoferrin-binding proteins with polylactosamine-binding ability; Eda S et al.; Human lactoferrin (LF) specifically binds to human monocytic leukemia cell line THP-1 cells differentiated into macrophages, and it has been suggested that the poly-N-acetyllactosaminyl saccharide chains of LF are involved . We partially purified and characterized LF-binding proteins with affinity for polylactosamines from THP-1 cells . LF-binding activity was solubilized by nonionic detergent Triton X-100 from THP-1 cell membrane, and subjected to affinity chromatography using an LF-Sepharose column . LF-binding activity, detected by ligand blotting assay, was eluted and further fractionated by affinity chromatography using a Sepharose column coupled with band 3, a polylactosaminyl chain-containing glycoprotein of human erythrocyte membrane . LF-binding activity was separated into three fractions (frs . B1, B2, and B3) . These fractions exhibited band 3-binding activity as detected by ligand blotting assay . Dodecylsulfate-polyacrylamide gel electrophoresis of frs . B1, B2, and B3, followed by detection of LF-binding activity on Western blots, indicated that frs . B1, B2, and B3 contained LF-binding proteins with a molecular mass of 35, 50 and 80, and 35-37 kDa, respectively . Binding of LF to each of the fractions on the dot blots was partially inhibited by LF oligosaccharides, band 3 oligosaccharides and lacto-N-neotetraose, each containing di-N-acetyllactosaminyl or analogous structure, Gal beta 1-->4GlcNAc beta 1-->3Gal beta 1-->4GlcNAc (or Glc) . These results suggest that the 35, 50 and/or 80, and 35-37 kDa proteins on THP-1 cells are LF-binding proteins with polylactosamine-binding ability. Biol Pharm Bull, 1997 Feb, 20(2), 122 - 6 Localization of enzymes involved in metabolism of glycyrrhizin in contents of rat gastrointestinal tract; Akao T; Most digested food 2 h after overnight feeding in rat remained in the stomach, duodenum, upper small intestine, lower small intestine, cecum and colon, all of which indicated pH between 4 and 7 and had glycyrrhizin (GL) hydrolyzing activity . This enzyme activity was highest in the cecal and colonic contents among all gastrointestinal contents . Also, 3 alpha-hydroxyglycyrrhetic acid (3 alpha-hydroxyGA) and 3 beta-hydroxyglycyrrhetic acid (3 beta-hydroxyGA) oxidizing enzymes were localized in the same cecal content . Namely, rat gastrointestinal bacteria had the ability to hydrolyze GL to 3 beta-hydroxyGA by glycyrrizin beta-D-glucuronidase and to oxide 3 beta-hydroxyGA and 3 alpha-hydroxyGA to 3-oxoGA by 3 beta-hydroxyglycyrrhetinate dehydrogenase and 3 alpha-hydroxyglycyrrhetinate dehydrogenase, respectively . In medium of pH 1 to pH 10, metabolites 3 beta-hydroxyGA, 3-oxoGA and 3 alpha-hydroxyGA obtained from the metabolism of GL were the highest in pH 8 . The intestinal contents of pH 6 or pH 7 were able to produce metabolites 3 beta-hydroxyGA in the metabolism of GL . However, the stomach content at pH 4.2 was lowest in metabolite 3 beta-hydroxyGA . It is unknown whether or not GL is metabolized to 3 beta-hydroxyGA by the stomach content in vivo. Am J Infect Control, 1997 Feb, 25(1), 11 - 5 A randomized trial of surgical scrubbing with a brush compared to antiseptic soap alone; Loeb MB et al.; BACKGROUND: The difference between use of a scrub brush versus soap alone in reducing hand bacterial counts has never been established by a prospective, comparative study . METHODS: Fifteen volunteers were taught the 5-minute surgical scrub . Baseline specimens were obtained by the glove fluid sampling procedure . Subjects were randomized to (1) scrub with an inert scrub brush and 4% chlorhexidine soap with isopropyl alcohol or (2) wash with 4% chlorhexidine soap with isopropyl alcohol alone . Specimens were obtained immediately after the scrub was completed and 45 minutes later . The experiment was repeated by use of a cross-over design after a 1-week washout period . The data were analyzed by three methods that took into account the broad range of baseline hand counts (5 x 10(1) to 11.2 x 10(4): method 1, the discordance between presence/absence of hand bacterial counts within individuals at 45 minutes for soap versus soap and brush; method 2, the absolute reduction of bacteria (baseline vs 45 min.) for soap versus soap and brush; and method 3, the proportional change in bacterial counts at 45 minutes from baseline for soap versus soap and brush . RESULTS: Although there was no statistically significant difference for any method, the point estimates for the odds ratio (OR) showed that up to twice the number of subjects had a greater reduction in bacterial counts when they washed with soap than when they scrubbed with a brush, as evidenced by the following data: method 1, OR 2.3 (95% confidence interval {CI} 0.53, 13.99) for soap alone; method 2, OR 1.0 (CI 0.23, 4.35); and method 3, OR 2.0 (CI 0.54, 9.10) for soap alone . CONCLUSIONS: The effect of use of soap alone in reducing hand bacterial counts at 45 minutes was similar to use of soap and brush . Soap can be used alone and the surgical infection rate prospectively monitored. Poult Sci, 1997 Feb, 76(2), 248 - 55 The applicability of particleboard residue as a litter material for male turkeys; Hester PY et al.; Particleboard residue is a by-product of the secondary wood products manufacturing industries . Large quantities of this product are landfilled for lack of better use . The objective of the current study was to investigate the possibility of using particleboard residue as a litter source for male turkeys . Two sizes of particleboard residue, fine and coarse, were compared to hardwood shavings . Compared to hardwood shavings, fine and coarse particleboard was a drier, cleaner product initially, as indicated by lower moisture content as well as bacteria and mold counts at Day 0 . Turkeys reared for 123 d on fine particleboard had several advantages over those reared on either the coarse particleboard or hardwood shavings, which included significantly lowered incidences of breast buttons and leg abnormalities . Perhaps due to the jagged edges and coarser texture, coarse particleboard increased the incidence of foot pad dermatitis when compared to the other two litter sources . Turkeys reared on fine particle-board had a 0.16 kg reduction (P < 0.01) in live market body weights compared to the toms reared on hardwood shavings, but this was offset by a 0.22 kg gain in muscle deposition (P < 0.05) . Mortality, breast weights and yields, and feed efficiency were unaffected by litter source . Based on the variables studied, it was concluded that fine particleboard residue could be used as an alternative bedding material for male turkeys. J Orthop Trauma, 1997 Feb-Mar, 11(2), 121 - 5 Benzalkonium chloride: a potential disinfecting irrigation solution; Gainor BJ et al.; OBJECTIVE: To determine the disinfecting properties of benzalkonium chloride as an irrigation agent . DESIGN: Comparison was made between irrigation of contaminated muscle strips with benzalkonium chloride and normal saline (control) . SUMMARY OF BACKGROUND DATA: Benzalkonium chloride is a cationic disinfectant, which has questionable efficacy in an organic environment . However, no previous study has attempted to use high volumes of this cationic solution to overcome the neutralizing effect of organic tissue and thus maintain this detergent's germicidal properties . METHODS: 2.5 cm x 0.5 cm x 0.5 cm pieces of bovine muscle were aseptically cut from the center of freshly harvested beef muscle and incubated with 1.0 x 10(7) colony forming units of bacteria for 15 minutes . The muscle strips were then irrigated with either 100 mL, 1 L, or 10 L of benzalkonium chloride at a 1:2000 concentration in normal saline . Normal saline was used as the control . The muscle strips were sonicated to remove adherent bacteria; the number of living organisms was determined by quantitatively culturing the sonicate . RESULTS: In vitro, benzalkonium chloride was superior to normal saline at disinfecting bovine muscle (p < or = 0.001) . When 10 L of benzalkonium chloride irrigation was used, no living bacteria could be recovered (p < or = 0.012) . CONCLUSION: In this experimental setting benzalkonium chloride was an effective disinfection agent, with enhanced activity at large volumes. J Anim Sci, 1997 Feb, 75(2), 556 - 65 Cellular defense mechanisms in the udder and lactation of goats; Paape MJ et al.; Migration of neutrophils into mammary tissue provides the first immunological line of defense against bacteria that penetrate the physical barrier of the teat canal . Evasion of neutrophil defenses provides an opportunity for invading bacteria to become established . Depletion of neutrophils results in a dramatic increase in susceptibility to intramammary infection . Numerous cytoplasmic particles are shed from the apical surface of mammary secretory cells during milk secretion in goats . Only those counting methods that are specific for deoxyribonucleic acid can distinguish cell-like particles from somatic cells and thereby give reliable estimates of somatic cell numbers in goat milk . Unlike in milk from dairy cows, the somatic cell count in goat milk is influenced by the presence of nucleated cytoplasmic particles, stage of lactation, parity, and caprine arthritis-encephalitis . Investigations indicate that a dry period is necessary for optimal milk production in dairy cows but may not be necessary in goats . However, in many other respects regulation of bovine and caprine lactation seems to be quite similar . Studies have demonstrated additive galactopoietic effects of growth hormone and frequent milking in both species and a recently isolated chemical feedback inhibitor of lactation seems effective across both species . Increasing lactational performance has the potential for decreasing milk somatic cell counts in late lactation. Naunyn Schmiedebergs Arch Pharmacol, 1997 Feb, 355(2), 190 - 7 In U-937 promonocytes, misteltoe lectin I increases basal {Ca2+}i, enhances histamine H1- and complement C5a-receptor-mediated rises in {Ca2+}i, and induces cell death; Wenzel-Seifert K et al.; Mistletoe lectin I (ML I) from Viscum album inhibits cell growth and induces apoptosis (programmed cell death) in several cell types . Because increases in cytosolic Ca2+ concentration ({Ca2+}i) constitute a signal for the induction of apoptosis, we studied the effects of ML I on basal {Ca2+}i, receptor-mediated rises in {Ca2+}i and cell viability, using human U-937 promonocytes as model system . Treatment of U-937 cells with ML I (30-100 ng/ml) significantly increased basal {Ca2+}i . ML I (10-30 ng/ml) enhanced histamine-induced rises in {Ca2+}i up to five-fold . The effect of histamine was inhibited by clemastine but not by famotidine, indicative for its mediation via H1-receptors . ML I additionally enhanced the stimulatory effect of complement C5a on {Ca2+}i, whereas the effect of ATP was unaffected . ML I did not induce responsiveness of U-937 cells towards a bacteria-derived chemotactic peptide . ML I up to 10 ng/ml did not affect cell viability and growth of U-937 cells . ML I at 30 ng/ml moderately inhibited cell growth and reduced cell viability . At 100 ng/ml, ML I was strongly cytotoxic . Our data support the view that Ca2+ plays a role as intracellular signal molecule in the induction of apoptosis and point to an accelerating role of H1- and C5a-receptors in the regulation of this process. Plant Mol Biol, 1997 Feb, 33(3), 381 - 92 Light induces accumulation of isocitrate lyase mRNA in a carotenoid-deficient mutant of Chlamydomonas reinhardtii; Petridou S et al.; A cDNA with sequence similarity to isocitrate lyase (ICL) genes was isolated from the unicellular eukaryotic green alga Chlamydomonas reinhardtii as a light-induced mRNA in the carotenoid biosynthetic mutant strain FN68 . The 416 amino acid open reading frame shows significant sequence similarity to isocitrate lyases of bacteria (70%), molds (48%), yeasts (45%), and plants (47%) . Expression of the Chlamydomonas ICL gene was tested in the mutant strain FN68, which when grown in the dark fails to accumulate carotenoids and is deficient in chlorophyll, and in CC400G, a strain that accumulates wild-type levels of carotenoids and chlorophyll . In vegetative CC400G cells, ICL mRNA accumulated to a high level in the dark and declined to a barely detectable level within 30 min of exposure to light . This response was more sensitive to white (tungsten filament) or red light than green or blue light, excluding cryptochrome and rhodopsin as the photoreceptor . These results are consistent with excitation by chlorophyll and/or a phytochrome-related photoreceptor . In vegetative FN68 cells, ICL mRNA abundance was very low in the dark, but increased dramatically in response to light . At intensities above threshold, excitation by far-red or red light-induced ICL mRNA accumulation to the highest levels . The threshold of the response was lowest for far-red and blue light . These results are consistent with excitation of a photochromic far-red-responsive pigment. Antonie Van Leeuwenhoek, 1997 Feb, 71(1-2), 159 - 78 Evolution of novel metabolic pathways for the degradation of chloroaromatic compounds; van der Meer JR; Chlorobenzenes are substrates not easily metabolized by existing bacteria in the environment . Specific strains, however, have been isolated from polluted environments or in laboratory selection procedures that use chlorobenzenes as their sole carbon and energy source . Genetic analysis indicated that these bacteria have acquired a novel combination of previously existing genes . One of these gene clusters contains the genes for an aromatic ring dioxygenase and a dihydrodiol dehydrogenase . The other contains the genes for a chlorocatechol oxidative pathway . Comparison of such gene clusters with those from other aromatics degrading bacteria reveals that this process of recombining or assembly of existing genetic material must have occurred in many of them . Similarities of gene functions between pathways suggest the incorporation of existing genetic material has been the most important mechanism of expanding a metabolic pathway . Only in a few cases a horizontal expansion, that is acquisition of gene functions to accommodate a wider range of substrates which are then all transformed in one central pathway, is observed on the genetic level . Evidence is presented indicating that the assembly process may trigger a faster divergence of nearby gene sequences . Further 'fine-tuning', for example by developing a proper regulation, is then the next step in the adaptation. Plant Physiol, 1997 Feb, 113(2), 463 - 8 Origin of the thiazole ring of camalexin, a phytoalexin from Arabidopsis thaliana; Zook M et al.; The principal phytoalexin that accumulates in Arabidopsis thaliana after infection by fungi or bacteria is 3-thiazol-2'-yl-indole (camalexin) . Detached noninoculated leaves of Arabidopsis and leaves inoculated with the fungus Cochliobolus carbonum were fed {35S}cysteine (Cys) and {35S}methionine . Inoculated leaves incorporated more than a 200-fold greater amount of radioactivity from {35S}Cys into camalexin, as compared with noninoculated leaves . The amount of radioactivity from {35S}Cys that was incorporated into camalexin from inoculated Arabidopsis leaves was 10-fold greater than the amount of radioactivity that was incorporated into camalexin from {35S}methionine . Additional labeling experiments were performed to determine whether other atoms of Cys are incorporated into camalexin . {14C}Cys and {35S}Cys were incorporated into camalexin with approximately the same efficiency . Cys labeled either with deuterium (D3-Cys{2,3,3}) or 13C and 15N ({U-13C,15N}Cys) was also fed to inoculated leaves of Arabidopsis; camalexin was analyzed by mass spectroscopic analysis . The average ratio of molecular ion intensities of 203/200 for {U-13C,15N}Cys-labeled camalexin was 4.22, as compared with 0.607 for the average 203/200 ratio for unlabeled camalexin . The mass fragment-ion intensity ratios of 60/58 (thiazole ring ion fragment) and 143/142 were also higher for {U-13C,15N}Cys-labeled camalexin, as compared with unlabeled camalexin . The 59/58 and 201/200 ratios were higher for D3-Cys-labeled camalexin as compared with unlabeled camalexin . These data are consistent with the predicted formation of the thiazole ring of camalexin from Cys. Bioessays, 1997 Feb, 19(2), 117 - 26 Chloride channels: an emerging molecular picture; Jentsch TJ et al.; Chloride channels are probably found in every cell, from bacteria to mammals . Their physiological tasks range from cell volume regulation to stabilization of the membrane potential, signal transduction, transepithelial transport and acidification of intracellular organelles . These different functions require the presence of many distinct chloride channels, which are differentially expressed and regulated by various stimuli . These include various intracellular messengers (like calcium and cyclic AMP), pH, extracellular ligands and transmembrane voltage . Three major structural classes of chloride channels are known to date, but there may be others not yet identified . After an overview of the general functions of chloride channels, this review will focus on these cloned chloride channels: the CLC chloride channel family, which includes voltage-gated chloride channels, and the cystic fibrosis transmembrane regulator (CFTR), which performs other functions in addition to being a chloride channel . Finally, a short section deals with GABA and glycine receptors . Diseases resulting from chloride channel defects will be specially emphasized, together with the somewhat limited information about how these proteins work at the molecular level. Mol Microbiol, 1997 Feb, 23(3), 537 - 49 Substrate induction and glucose repression of maltose utilization by Streptomyces coelicolor A3(2) is controlled by malR, a member of the lacl-galR family of regulatory genes; van Wezel GP et al.; malR of Streptomyces coelicolor A3(2) encodes a homologue of the Lacl/GalR family of repressor proteins, and is divergently transcribed from the malEFG gene cluster, which encodes components of an ATP-dependent transport system that is required for maltose utilization . Transcription of malE was induced by maltose and repressed by glucose . Disruption or deletion of malR resulted in constitutive, glucose-insensitive malE transcription at a level markedly above that observed in the parental malR+ strain, and overproduction of MalR prevented growth on maltose as carbon source . Consequently, MalR plays a crucial role in both substrate induction and glucose repression of maltose utilization . malR is expressed from a single promoter with transcription initiating at the first G of the predicted GTG translation start codon. Microbiology, 1997 Feb, 143 ( Pt 2), 539 - 46 A new insertion sequence IS1452 from Acetobacter pasteurianus; Kondo K et al.; A new insertion sequence element, IS1452, was found to be associated with inactivation of the alcohol dehydrogenase by insertion in the adhS gene encoding subunit III of the three-component membrane-bound alcohol dehydrogenase complex in Acetobacter pasteurianus . Cloning and sequencing analyses of the mutated subunit III gene locus revealed that IS1452 was inserted at or near the ribosome-binding sequence of adhS . Analysis of transcription using the chloramphenicol acetyltransferase gene as the reporter indicated that IS1452 abolished transcription of adhS by separating its promoter from the subunit III structural gene . IS1452 was 1411 bp in length and had a terminal inverted repeat of 21 bp . IS1452 contained one long ORF of 416 amino acids rich in basic amino acids . This protein showed homology with a putative transposes, Tra1, of IS701 isolated from the cyanobacterium Calothrix species PCC 7601 . Like IS701, IS1452 was found to generate a 4 bp direct repeat at the site of insertion upon transposition . The target site specificity was rather strict, and a CTA(A or G) sequence appeared to be preferentially recognized . Transposition of IS1452 was replicative, since it was accompanied by an increase in the copy number of IS1452 . Several strains belonging to the genus Acetobacter also contained IS1452 at varying copy numbers from one to more than ten . These observations suggest that IS1452 is one of the insertion sequences that are responsible for genetic instability leading to deficiencies in various physiological properties in acetic acid bacteria. Scand J Immunol, 1997 Feb, 45(2), 115 - 31 Host responses and antigens involved in protective immunity to Mycobacterium tuberculosis; Andersen P; Tuberculosis (TB) is the largest single infectious cause of human mortality . The incidence of TB has remained high in most of the developing world and the disease has recently re-emerged as a public health problem in industrialized countries . The development of a new improved TB vaccine is a highly prioritized international research area, which has been further stimulated by the appearance of multi-drug resistant strains of Mycobacterium tuberculosis . The present status of the attempts to characterize the protective immune response to TB will be reviewed with special emphasis on recent progress in the identification and characterization of target molecules recognized by protective cells . This paper will focus on proteins released from live bacteria and discuss their role in the host-pathogen interaction and the ongoing attempts to use these molecules in TB subunit vaccines. Protein Sci, 1997 Feb, 6(2), 450 - 8 Thermoanaerobacter brockii alcohol dehydrogenase: characterization of the active site metal and its ligand amino acids; Bogin O et al.; The active-site metal ion and the associated ligand amino acids in the NADP-linked, tetrameric enzyme Thermoanaerobacter brockii alcohol dehydrogenase (TBADH) were characterized by atomic absorption spectroscopy analysis and site-directed mutagenesis . Our preliminary results indicating the presence of a catalytic zinc and the absence of a structural metal ion in TBADH (Peretz & Burstein . 1989 . Biochemistry 28:6549-6555) were verified . To determine the role of the putative active-site zinc, we investigated whether exchanging the zinc for other metal ions would affect the structural and/or the enzymatic properties of the enzyme . Substituting various metal ions for zinc either enhanced or diminished enzymatic activity, as follows: Mn2+ (240%); Co2+ (130%); Cd2+ (20%); Cu2+ or V3+ (< 5%) . Site-directed mutagenesis to replace any one of the three putative zinc ligands of TBADH, Cys 37, His 59, or Asp 150, with the non-chelating residue, alanine, abolished not only the metal-binding capacity of the enzyme but also its catalytic activity, without affecting the overall secondary structure of the enzyme . Replacing the three putative catalytic zinc ligands of TBADH with the respective chelating residues serine, glutamine, or cysteine damaged the zinc-binding capacity of the mutated enzyme and resulted in a loss of catalytic activity that was partially restored by adding excess zinc to the reaction . The results imply that the zinc atom in TBADH is catalytic rather than structural and verify the involvement of Cys 37, His 59, and Asp 150 of TBADH in zinc coordination. J Bone Miner Res, 1997 Feb, 12(2), 165 - 71 Human stanniocalcin inhibits renal phosphate excretion in the rat; Wagner GF et al.; Stanniocalcin (STC) is a glycoprotein hormone first identified in bony fishes where it counteracts hypercalcemia by inhibiting gill calcium uptake and stimulating renal inorganic phosphate (Pi) reabsorption . Human STC (hSTC) has recently been cloned and sequenced and is highly homologous to the fish hormone at the amino acid level . The objective of this study was to examine the possible effects of hSTC on electrolyte homeostasis and renal function in the rat . Recombinant hSTC was expressed in bacteria and purified by metal-ion affinity chromatography and reverse-phase high performance liquid chromatography . Anesthetized animals were given bolus infusions of 1, 5, or 10 nmol hSTC per kilogram of body weight . Control animals received solvent alone . The most effective dosage was 5 nmol/kg, which caused significant reductions in both absolute and fractional phosphate excretion in comparison with control rats . The hSTC had no effect on the renal excretion of other ions, the glomerular filtration rate, renal blood flow, blood pressure, or plasma electrolytes (Na+, K+, Ca2+, Pi, Mg/+) . The maximum effect of hSTC on phosphate excretion was observed 60-80 minutes postinjection . Lesser effects were obtained with higher and lower dosages of hormone . When renal cortical brush-border membrane vesicles were isolated from control and hormone-treated animals 80 minutes postinjection, the rate of Na+/Pi cotransport was found to be 40% higher in vesicles from hormone-treated animals (p < 0.01; 5 nmol hSTC/kg) . Together, the renal clearance and membrane vesicle data indicate that hSTC participates in the renal regulation of Pi homeostasis in mammals. Curr Opin Immunol, 1997 Feb, 9(1), 17 - 23 Cytokines acting on or secreted by macrophages during intracellular infection (IL-10, IL-12, IFN-gamma); Trinchieri G; The three cytokines IL-12, IL-10, and IFN-gamma have important and cross-regulatory roles in infection . In the past year, much progress has been made in the understanding of the cellular and molecular mechanisms involved in the regulation (and cross-regulation) of these three cytokines and their role in pathology . IL-12 is rapidly produced after infection and acts as a proinflammatory cytokine eliciting the production, by T cells and natural killer cells, of IFN-gamma which activates phagocytic cells . The production of IL-12 is strictly regulated by negative and positive feedback mechanisms . If IL-12 and IL-12-induced IFN-gamma are present during early T cell expansion in response to antigen, Th1 cell generation is favored and the generation of Th2 cells is inhibited . Thus, IL-12 is also a potent immunoregulatory cytokine which promotes Th1 differentiation and is instrumental in the Th1-dependent resistance to infections by bacteria, intracellular parasites, fungi, and certain viruses . Viruses inducing a permanent or transient immunodepression, such as HIV and measles, may act, in part, by suppressing IL-12 production. Nat Struct Biol, 1997 Feb, 4(2), 140 - 6 The structure of the cytochrome p450BM-3 haem domain complexed with the fatty acid substrate, palmitoleic acid; Li H et al.; The substrate-bound structures of two cytochrome P450s, P450cam and P450eryF, are known . While these structures reveal important features that control substrate specificity, the problem of how conformational changes allow for substrate entry and product release remains unsolved . The structure of the haem domain of the bacterial fatty acid hydroxylase, P450BM-3, previously was solved in the substrate-free form . Unlike the substrate-bound P450cam and P450eryF structures, the substrate access channel is open in substrate-free P450BM-3 . Here we present the X-ray structure of P450BM-3 at 2.7 A bound with a fatty acid substrate, palmitoleic acid . A comparison of the substrate-bound and -free forms reveals major conformational differences and provides the first detailed picture of substrate-induced conformational changes in a P450. Appl Environ Microbiol, 1997 Feb, 63(2), 687 - 93 Transformation yields of chlorinated ethenes by a methanotrophic mixed culture expressing particulate methane monooxygenase; Anderson JE et al.; Transformation yields for the aerobic cometabolic degradation of five chlorinated ethenes were determined by using a methanotrophic mixed culture expressing particulate methane monooxygenase (pMMO) . Transformation yields (expressed as moles of chlorinated ethene degraded per mole of methane consumed) were 0.57, 0.25, 0.058, 0.0019, and 0.00022 for trans-1,2-dichloroethylene (t-DCE), vinyl chloride (VC), cis-1,2-dichloroethylene (c-DCE), trichloroethylene (TCE), and 1,1-dichloroethylene (1,1-DCE), respectively . Degradation of t-DCE and VC was observed only in the presence of formate or methane, sources of reducing energy necessary for cometabolism . The t-DCE and VC transformation yields represented 35 and 15%, respectively, of the theoretical maximum yields, based on reducing-energy availability from methane dissimilation to carbon dioxide, exclusive of all other processes that require reducing energy . The yields for t-DCE and VC were 20 times greater than the yields reported by others for cells expressing soluble methane monooxygenase (sMMO) . Transformation yields for c-DCE, TCE, and 1,1-DCE were similar to or less than those for cultures expressing sMMO . Although methanotrophic biotreatment systems have typically been designed to incorporate cultures expressing sMMO, these results suggest that pMMO expression may be highly advantageous for degradation of t-DCE or VC . It may also be much easier to maintain pMMO expression in treatment systems, because pMMO is expressed by all methanotrophs whereas sMMO is expressed only by type II methanotrophs under copper-limited conditions. J Bacteriol, 1997 Feb, 179(4), 1143 - 52 Cloning and sequence of cymA, a gene encoding a tetraheme cytochrome c required for reduction of iron(III), fumarate, and nitrate by Shewanella putrefaciens MR-1; Myers CR et al.; The cymA gene, which encodes a tetraheme cytochrome c, was cloned from Shewanella putrefaciens MR-1 . This gene complemented a mutant which had a TnphoA insertion in cymA and which was deficient in the respiratory reduction of iron(III), nitrate, fumarate, and manganese(IV) . The 561-bp nucleotide sequence of cymA encodes a protein of 187 amino acids with a predicted molecular mass of 20.8 kDa . No N-terminal signal sequence was readily apparent; consistent with this, a cytochrome with a size of 21 kDa was detected in the wild type but was absent in the insertional mutant . The cymA gene is transcribed into an mRNA; the major transcript was approximately 790 bases, suggesting that it is not part of a multicistronic operon . This RNA transcript was not detected in the cymA mutant . The CymA protein was found in the cytoplasmic membrane and soluble fraction of MR-1, and it shares partial amino acid sequence homology with multiheme c-type cytochromes from other bacteria . These cytochromes are ostensibly involved in the transfer of electrons from the cytoplasmic membrane to acceptors in the periplasm . The localization of the fumarate and iron(III) reductases to the periplasm and outer membrane of MR-1, respectively, suggests the possibility of a similar electron transfer role for CymA. J Leukoc Biol, 1997 Feb, 61(2), 133 - 40 Marked inhibition of the intracellular multiplication of Legionella pneumophila in monocytes isolated from carriers of human T lymphotrophic virus type I; Funai N et al.; Intracellular growth patterns of Legionella pneumophila were examined in monocytes obtained from carriers of human T-lymphotropic virus type I (HTLV-1) and controls who were HTLV-1 seronegative . All subjects were seronegative for antibodies against L . pneumophila . Bacterial growth was determined 0, 1, 2, and/or 3 days after infecting peripheral blood mononuclear cells (PBMCs) with the bacteria . The intracellular growth of L . pneumophila was markedly inhibited in HTLV-1 carriers compared with normal controls . When the lymphocytes were depleted from the HTLV-1 carrier PBMC cultures before infection, this inhibition was abolished . Inhibition reappeared, however, when the 72-h culture supernatants of PBMCs from HTLV-1 carriers were added to the lymphocyte-depleted cultures . Culture supernatants of infected and uninfected PBMCs from HTLV-1 carriers exhibited markedly increased levels of interferon-gamma (IFN-gamma) and tumor necrosis factor alpha (TNF-alpha) compared with the HTLV-1 seronegative controls . In the HTLV-1 carriers, IFN-gamma was produced by the CD4+ lymphocytes, whereas TNF-alpha was secreted by the monocytes . Addition of anti-IFN-gamma or anti-TNF-alpha antibodies to the HTLV-1 carrier PBMC cultures diminished the inhibition of intracellular growth of L . pneumophila . Results suggest that the monocytes are activated in HTLV-1 carriers . These findings may explain why an opportunistic infection by certain intracellular pathogens is rarely seen among HTLV-1 carriers. Antimicrob Agents Chemother, 1997 Feb, 41(2), 475 - 7 Comparative evaluation of the inhibitory activities of the novel penicillanic acid sulfone Ro 48-1220 against beta-lactamases that belong to groups 1, 2b, and 2be; Tzouvelekis LS et al.; The inhibitory activity of the penicillanic acid sulfone Ro 48-1220 against group 1, 2b, and 2be beta-lactamases was evaluated . Ro 48-1220 inhibited TEM and SHV as effectively as clavulanate and tazobactam . It also inhibited group 1 beta-lactamases at lower concentrations than tazobactam . Ro 48-1220, at a concentration of 4 micrograms/ml, protected ceftriaxone and ceftazidime against strains producing group 1 and 2be beta-lactamases. Antimicrob Agents Chemother, 1997 Feb, 41(2), 379 - 84 A cytotoxicity assay for evaluation of candidate anti-Pneumocystis carinii agents; Cushion MT et al.; A series of over 60 agents representing several different classes of compounds were evaluated for their effects on the ATP pools of Pneumocystis carinii populations derived from immunosuppressed rats . A cytotoxicity assay based on an ATP-driven bioluminescent reaction was used to determine the concentration of agent which decreased the P . carinii ATP pools by 50% versus untreated controls (IC50) . A ranking system based on the IC50 value was devised for comparison of relative responses among the compounds evaluated in the cytotoxic assay and for comparison to in vivo efficacy . With few exceptions, there was a strong correlation between results from the ATP assay and the performance of the compound in vivo . Antibiotics, with the exception of trimethoprim-sulfamethoxazole (TMP-SMX), were ineffective at reducing the ATP pools and were not active clinically or in the rat model of P . carinii pneumonia . Likewise, other agents not expected to be effective, e.g., antiviral compounds, did not show activity . Standard anti-P . carinii compounds, e.g., TMP-SMX, pentamidine, and dapsone, dramatically reduced ATP levels . Analogs of the quinone and topoisomerase inhibitor groups were shown to reduce ATP concentrations and hold promise for further in vivo investigation . The cytotoxicity assay provides a rapid assessment of response, does not rely on replicating organisms, and should be useful for assessment of structure-function relationships. Am J Clin Oncol, 1997 Feb, 20(1), 24 - 30 Phase I trial of etoposide, carboplatin, and GM-CSF in extensive small-cell lung cancer: a Cancer and Leukemia Group B study (CALGB 8832); Luikart SD et al.; The maximum tolerated dose (MTD) of etoposide and carboplatin without growth factor support was previously defined by Cancer and Leukemia Group B (CALGB) as 200 and 125 mg/m2/day x 3, respectively, given every 28 days to previously untreated patients who have extensive, small-cell lung cancer (SCLC) . Myelosuppression was dose-limiting . The purpose of this phase I trial was to determine if granulocyte macrophage colony-stimulating factor (GM-CSF) support allows the dosage of the combination of etoposide and carboplatin to be increased above the previously determined MTD . In this CALGB study of 44 evaluable patients with performance status 0-2, cohorts were treated with etoposide and carboplatin given intravenously on days 1-3 followed by GM-CSF (molgramostim) given subcutaneously on days 4-18 . Four dose levels of bacteria-derived recombinant GM-CSF (5, 10, 20 microg/kg/day and 5 microg/kg every 12 h), three dose levels of etoposide (200, 250, and 300 mg/m2/day x 3), and two dose levels of carboplatin (125 and 150 mg/m2/day x 3) were evaluated . There was no chemotherapy dose escalation in individual patients . With 5 microg/kg/d GM-CSF, the first etoposide and carboplatin cycle of 300 and 150 mg/m2/day x 3, respectively, could be administered with acceptable toxicity . However, GM-CSF did not allow repeated administration of this dose-escalated regimen every 21 days, since delayed platelet and/or neutrophil recovery was dose limiting in later cycles . These results demonstrate that GM-CSF alone has limited capability to support the repeated administration of high doses of etoposide and carboplatin . CALGB currently is testing the ability of interleukin (IL)-6 given with GM-CSF to ameliorate the cumulative myelosuppression of this intense regimen. Infect Immun, 1997 Feb, 65(2), 750 - 7 Inhibition of Helicobacter pylori binding to gastrointestinal epithelial cells by sialic acid-containing oligosaccharides; Simon PM et al.; Helicobacterpylori, the ulcer pathogen residing in the human stomach, binds to epithelial cells of the gastric antrum . We have examined binding of 13 bacterial isolates to epithelial cell lines by use of a sensitive microtiter plate method in which measurement of bacterial urease activity provides the means for quantitation of bound organisms . Several established human gastrointestinal carcinoma cell lines grown as monolayers were compared for suitability in these assays, and the duodenum-derived cell line HuTu-80 was selected for testing bacterial binding inhibitors . When bacteria are pretreated with oligosaccharides, glycoproteins, and glycolipids, a complex picture of bacterial-epithelial adherence specificities emerges . Among the monovalent inhibitors tested, 3'-sialyllactose (NeuAc alpha2-3Gal beta1-4Glc; 3'SL) was the most active oligosaccharide, inhibiting adherence for recent clinical isolates of H . pylori with a millimolar 50% inhibitory concentration (IC50) . Its alpha2-6 isomer (6'SL) was less active . Most of the recent clinical isolates examined were inhibited by sialyllactose, whereas long-passaged isolates were insensitive . Among the long-passaged bacterial strains whose binding was not inhibited by 3'SL was the strain ATCC 43504, also known as NCTC 11637 and CCUG 17874, in which the proposed sialyllactose adhesin was recently reported to lack surface expression (P . G . O'Toole, L . Janzon, P . Doig, J . Huang, M . Kostrzynska, and T . H . Trust, J . Bacteriol . 177:6049-6057, 1995) . Pretreatment of the epithelial monolayer with neuraminidase reduced the extent of binding by those bacteria that are sensitive to inhibition by 3'SL . Other potent inhibitors of bacterial binding are the glycoproteins alpha1-acid glycoprotein, fetuin, porcine gastric and bovine submaxillary mucins, and the glycolipid sulfatide, all of which present multivalent sialylated and/or sulfated galactosyl residues under the conditions of the binding assay . Consistent with this pattern, a multivalent neoglycoconjugate containing 20 mol of 3'SL per mol of human serum albumin inhibited bacterial binding with micromolar IC50 . The H . pylori isolate most sensitive to inhibition by 3'SL was least sensitive to inhibition by sulfatide, gastric mucin, and other sulfated oligosaccharides . Bacteria that have been allowed to bind epithelial cells are also effectively detached by 3'SL . These results describe a heterogeneous adherence repertoire for these bacteria, but they also confirm the critical role of the 3'SL structure on human gastric epithelial cells as an adherence ligand for recent isolates of H . pylori. Infect Immun, 1997 Feb, 65(2), 422 - 7 Role of the carboxyl-terminal region of Porphyromonas gingivalis fimbrillin in binding to salivary proteins; Nagata H et al.; Porphyromonas gingivalis fimbriae are considered to play an important role in the adherence and colonization of the bacteria in the oral cavity . In this study, we generated and purified three carboxyl-terminal variants of recombinant fimbrillin (r-FimA 224-337, r-FimA 266-337, and r-FimA 287-337, corresponding to amino acid residues 224 to 337, 266 to 337, and 287 to 337, respectively, of the 43-kDa fimbrillin of P . gingivalis 2561) . They were used as inhibitors of P . gingivalis cell binding to human salivary protein-coated hydroxyapatite (HAP) beads . All of the carboxyl-terminal region polypeptides inhibited binding in a dose-dependent manner; however, the inhibitory effect of r-FimA 287-337 was less than that of the other two polypeptides when HAP beads were coated with whole saliva or purified salivary proline-rich protein 1 (PRP1) . Assays of binding of a synthetic peptide corresponding to amino acid residues 266 to 286 of P . gingivalis 2561 fimbrillin to salivary proteins showed that this peptide bound strongly to whole saliva or PRP1 but only weakly to statherin . These results suggest that the carboxyl-terminal region corresponding to amino acid residues 266 to 337 of P . gingivalis fimbrillin plays an important role in binding to salivary proteins and that the domain corresponding to amino acids 266 to 286 is likely a major binding site for PRP1s and the domain corresponding to amino acids 287 to 337 is likely a major binding site for statherin. J Bacteriol, 1997 Feb, 179(3), 663 - 9 Analysis of the cbbXYZ operon in Rhodobacter sphaeroides; Gibson JL et al.; Three genes, cbbX, cbbY, and cbbZ were found downstream from the form I ribulose 1,5-bisphosphate carboxylase/oxygenase (RubisCO) genes of Rhodobacter sphaeroides . As in chemoautotrophic bacteria, cbbZ was shown to encode phosphoglycolate phosphatase (PGP), whereas the identities of cbbX and cbbY are not known . To determine the physiological function of the cbbXYZ gene products, we constructed R . sphaeroides strains in which the genes were inactivated and characterized the resultant mutant strains according to growth phenotype and levels of RubisCO and PGP . Only a mutation in cbbX resulted in a discernible phenotype, namely, impaired photoautotrophic growth . No PGP activity was observed in any of the mutants, suggesting that the three genes are transcriptionally linked . Studies with a spontaneous chemoautotrophic competent derivative of the CBBX mutant suggested that the cbbXYZ gene products are not essential for chemoautotrophic growth . PGP activity determined in the wild-type strain grown under a variety of growth conditions, and in various strains containing mutations in Calvin-Benson-Bassham cycle structural and regulatory genes, indicated that transcription of the cbb(I) operon influenced expression of the downstream cbbXYZ operon. J Clin Microbiol, 1997 Feb, 35(2), 455 - 61 Concordance of Porphyromonas gingivalis colonization in families; Tuite-McDonnell M et al.; Periodontitis is a widespread disease that appears to be due to a specific bacterial infection . Several species of bacteria have been investigated as potential pathogens, and particularly strong evidence links the presence of Porphyromonas gingivalis with indicators of periodontitis . Information concerning the transmission of P . gingivalis between human contacts may be important in determining risk factors for disease and developing preventive strategies . A few small studies have provided some evidence of transmission between related individuals, but no large-scale study of families that would reflect the typical transmission of this pathogen in the population has been reported . The purpose of this study was to investigate the transmission of P . gingivalis within randomly selected, extended families . The colonization status of 564 members of multigeneration families was determined, and the degree of concordance observed among members of these families was then compared to that expected to occur based on the prevalence of colonization in the population studied . A PCR assay was used for detection of P . gingivalis . Concordance in colonization was more frequently observed within entire families (P = 0.0000) and for spouses (P < 0.001), children and their mothers (P < 0.001), children and their fathers (P < 0.01), adults and their mothers (P < 0.005), and siblings (P < 0.05) than would be expected if P . gingivalis were randomly distributed in the population studied . Results showed that contact with an infected family member substantially increased the relative risk of colonization in these intrafamilial pairs . This indicates that P . gingivalis is commonly transmitted by contact with an infected family member. Endocrinology, 1997 Feb, 138(2), 602 - 6 Thyroid-stimulating hormone-induced down-regulation of thyroid transcription factor 1 in rat thyroid FRTL-5 cells; Saito T et al.; Thyroid transcription factor 1 (TTF-1) is thought to play an important role in the expression of genes that encode thyroid-specific proteins such as thyroglobulin, thyroid peroxidase, and TSH-receptor . The role of TSH in the regulation of TTF-1 messenger RNA (mRNA) and protein abundance was investigated in rat thyroid FRTL-5 cells . Northern blot analysis revealed that TSH reduced TTF-1 mRNA abundance in a dose- and time-dependent manner . Immunoblot analysis with rabbit antibodies prepared against a recombinant fragment of TTF-1 expressed in bacteria showed that TSH also reduced the amount of TTF-1 protein in FRTL-5 cells . Whereas the effect of TSH on TTF-1 mRNA was apparent after 3 h, the effect on TTF-1 protein was not apparent until 12 h after TSH addition to the cells . Both TTF-1 mRNA and protein were significantly decreased after the addition of (Bu)2 cAMP or forskolin for 24 h, whereas they were not decreased by 12-O-tetradecanoyl-phorbol-13-acetate . These results indicate that TSH down-regulates TTF-1 expression in FRTL-5 cells via the cAMP pathway. J Virol, 1997 Feb, 71(2), 1547 - 57 Physical and functional interactions between herpes simplex virus immediate-early proteins ICP4 and ICP27; Panagiotidis CA et al.; The ordered expression of herpes simplex virus type 1 (HSV-1) genes, during the course of a productive infection, requires the action of the virus immediate-early regulatory proteins . Using a protein interaction assay, we demonstrate specific in vitro protein-protein interactions between ICP4 and ICP27, two immediate-early proteins of HSV-1 that are essential for virus replication . We map multiple points of contact between these proteins . Furthermore, by coimmunoprecipitation experiments, we demonstrate the following . (i) ICP4-ICP27 complexes are present in extracts from HSV-1 infected cells . (ii) ICP27 binds preferentially to less modified forms of ICP4, a protein that is extensively modified posttranslationally . We also demonstrate, by performing electrophoretic mobility shift assays and supershifts with monoclonal antibodies to ICP4 or ICP27, that both proteins are present in a DNA-protein complex with a noncanonical ICP4 binding site present in the HSV thymidine kinase (TK) gene . ICP4, in extracts from cells infected with ICP27-deficient viruses, is impaired in its ability to form complexes with the TK site but not with the canonical site from the alpha4 gene . However, ICP4 is able to form complexes with the TK probe, in the absence of ICP27, when overproduced in mammalian cells or expressed in bacteria . These data suggest that the inability of ICP4 from infected cell extracts to bind the TK probe in the absence of ICP27 does not reflect a requirement for the physical presence of ICP27 in the complex . Rather, they imply that ICP27 is likely to modulate the DNA binding activity of ICP4 by affecting its posttranslational modification status . Therefore, we propose that ICP27, in addition to its established role as a posttranscriptional regulator of virus gene expression, may also modulate transcription either through direct or indirect interactions with HSV regulatory regions, or through its ability to modulate the DNA binding activity of ICP4. J Biol Chem, 1997 Jan 31, 272(5), 2936 - 41 Cloning and expression of a novel mammalian thioredoxin; Spyrou G et al.; We have isolated a 1276-base pair cDNA from a rat heart cDNA library that encodes a novel thioredoxin (Trx2) of 166 amino acid residues with a calculated molecular mass of 18.2 kDa . Trx2 possesses the conserved thioredoxin-active site, Trp-Cys-Gly-Pro-Cys, but lacks structural cysteines present in all mammalian thioredoxins . Trx2 also differs from the previously described rat thioredoxin (Trx1) by the presence of a 60-amino acid extension at the N terminus . This extension has properties characteristic for a mitochondrial translocation signal, and the cleavage at a putative mitochondrial peptidase cleavage site would give a mature protein of 12.2 kDa . Western blot analysis from cytosolic, peroxisomal, and mitochondrial rat liver cell fractions confirmed mitochondrial localization of Trx2 . Northern blot and reverse transcriptase-polymerase chain reaction analyses revealed that Trx2 hybridized to a 1.3-kilobase message, and it was expressed in several tissues with the highest expression levels in heart, muscle, kidney, and adrenal gland . N-terminally truncated recombinant protein was expressed in bacteria and characterized biochemically . Trx2 possessed a dithiol-reducing enzymatic activity and, with mammalian thioredoxin reductase and NADPH, was able to reduce the interchain disulfide bridges of insulin . Furthermore, Trx2 was more resistant to oxidation than Trx1. J Biol Chem, 1997 Jan 31, 272(5), 2914 - 9 Identification of the region in actin-binding protein that binds to the cytoplasmic domain of glycoprotein IBalpha; Meyer SC et al.; Actin-binding protein (ABP-280) is a component of the submembranous cytoskeleton and interacts with the glycoprotein (GP) Ibalpha subunit of the GP Ib-IX complex in platelets . In the present studies, we have identified the binding site for GP Ibalpha in ABP-280 . A melanoma cell line lacking ABP-280 was stably transfected with the cDNAs coding for GP Ib-IX, then transiently transfected with cDNA coding for various carboxyl-truncates of ABP-280 . Immunocapture assays and co-immunoprecipitation experiments from detergent-lysed cells showed that deletion of the carboxyl-terminal repeats 20-24 of ABP-280 had no effect on GP Ib-IX binding, but deletion of residues 2099 through 2136 within repeat 19 abolished binding . In the yeast two-hybrid system, an ABP-280 fragment comprising repeats 17-19 bound GP Ibalpha . Deletion from either end abolished binding . Individual or multiple repeats of ABP-280 were expressed as fusion protein in bacteria and purified; structural folding was evaluated, and binding to GP Ib-IX was assessed . Binding depended on the presence of repeats 17-19 . None of the individual repeats were able to bind to GP Ib-IX . These findings demonstrate that residues 1850-2136 comprising repeats 17-19 contain the binding site for GP Ib-IX. Fortschr Med, 1997 Jan 30, 115(3), 35 - 6 {Flies--reservoirs and vectors of Helicobacter pylori}; Grubel P et al.; The route by which humans get infected with H.p . is unknown . Viable bacteria have been shown to be excreted in human feces from infected individuals . We have recently shown that flies are able to carry and disseminate viable H . p . with their excreta, when they have fed on H.p . We propose a hypothesis that H . p . is acquired from human feces by flies, which then, while crawling on human food, contaminate it with their intestinal excreta . The food, swallowed by a susceptible individual then leads to a new infection . Flies should be considered prime suspects for the transmission of H . p., particularly in developing countries, where sanitary and domestic facilities are poor. FEBS Lett, 1997 Jan 27, 402(1), 50 - 2 Expression and activity of a recombinant chimeric protein composed of pokeweed antiviral protein and of human interleukin-2; Dore JM et al.; The pokeweed antiviral protein (PAP) has already been used to chemically construct immunotoxins . Here we tested the recombinant approach for the production of PAP-containing cytotoxic fusion-proteins . A cDNA encoding a mutated PAP (PAP9), which is expressed at high levels in bacteria, was fused to human interleukin-2 (IL-2) cDNA . The resulting PAP9-IL-2 protein was as active as the free PAP9 in inhibiting an eukaryotic cell-free translation system . Only the chimeric protein desaminated the 28S rRNA and inhibited translation of the CTLL-2 cell line which expresses the IL-2 receptor . These results show that PAP is a suitable toxin for the production of recombinant immunotoxins. J Biol Chem, 1997 Jan 24, 272(4), 2129 - 35 Evaluation of secondary structure of OxlT, the oxalate transporter of Oxalobacter formigenes, by circular dichroism spectroscopy; Fu D et al.; OxlT, the oxalate/formate exchange transporter of Oxalobacter formigenes, was purified as a histidine-tagged variant, OxlTHis, using Ni2+-linked affinity chromatography . OxlTHis was readily obtained in high purity (>/=95%) and reasonable yield (>/=60%), and showed kinetic and biochemical features characteristic of its parent, OxlT, including an unusually high maximal velocity (60 micromol/min per mg of protein at 4 degrees C) . Circular dichroism spectroscopy of purified OxlTHis identified the alpha-helix as its dominant secondary structural unit, encompassing 60-70% of OxlTHis residues and consistent with a model suggesting 60% of OxlT (OxlTHis) residues are involved in the construction of 12 transmembrane alpha-helices (Abe, K., Ruan, Z.-S., and Maloney, P . C . (1996) J . Biol . Chem . 271, 6789-6793) . In either octyl glucoside/lipid or dodecylmaltoside/lipid micelles, solubilized OxlTHis showed a striking substrate-induced stabilization of function, and at saturating levels of substrate (1000 x KD) activity recoverable by reconstitution disappeared with a half-life of 7 days at 23 degrees C . Measurement of changes of ellipticity at 222 nm as a function of time and substrate concentration showed that maintenance of function was attributable to a substrate-induced stabilization of the alpha-helical ensemble with a KD of 10 microM for the 1:1 binding of oxalate to OxlTHis. Science, 1997 Jan 24, 275(5299), 515 - 8 An Fe2IVO2 diamond core structure for the key intermediate Q of methane monooxygenase; Shu L et al.; A new paradigm for oxygen activation is required for enzymes such as methane monooxygenase (MMO), for which catalysis depends on a nonheme diiron center instead of the more familiar Fe-porphyrin cofactor . On the basis of precedents from synthetic diiron complexes, a high-valent Fe2(micro-O)2 diamond core has been proposed as the key oxidizing species for MMO and other nonheme diiron enzymes such as ribonucleotide reductase and fatty acid desaturase . The presence of a single short Fe-O bond (1.77 angstroms) per Fe atom and an Fe-Fe distance of 2.46 angstroms in MMO reaction intermediate Q, obtained from extended x-ray absorption fine structure and Mossbauer analysis, provides spectroscopic evidence that the diiron center in Q has an Fe2IVO2 diamond core. Biochemistry, 1997 Jan 21, 36(3), 489 - 94 Three-dimensional structure of tetrahydrodipicolinate N-succinyltransferase; Beaman TW et al.; The conversion of tetrahydrodipicolinate and succinyl-CoA to N-succinyltetrahydrodipicolinate and CoA is catalyzed by tetrahydrodipicolinate N-succinyltransferase and is the committed step in the succinylase pathway by which bacteria synthesize L-lysine and meso-diaminopimelate, a component of peptidoglycan . The X-ray crystal structure of THDP succinyltransferase has been determined to 2.2 A resolution and has been refined to a crystallographic R-factor of 17.0% . The enzyme is trimeric and displays the left-handed parallel beta-helix (L beta H) structural motif encoded by the "hexapeptide repeat" amino acid sequence motif {Raetz, C.R.H., & Roderick, S.L . (1995) Science 270, 997-1000} . The approximate location of the active site of THDP succinyltransferase is suggested by the proximity of binding sites for two inhibitors: p-(chloromercuri)benzenesulfonic acid and cobalt ion, both of which bind to the L beta H domain. J Exp Med, 1997 Jan 20, 185(2), 317 - 28 Maturation stages of mouse dendritic cells in growth factor-dependent long-term cultures; Winzler C et al.; The signals controlling the checkpoints of dendritic cells (DC) maturation and the correlation between phenotypical and functional maturational stages were investigated in a defined model system of growth factor-dependent immature mouse DC . Three sequential stages of DC maturation (immature, mature, and apoptotic) were defined and characterized . Immature DC (stage 1) had low expression of costimulatory molecules, highly organized cytoskeleton, focal adhesion plaques, and slow motility; accordingly, they were very efficient in antigen uptake and processing of soluble proteins . Further, at this stage most of major histocompatibility complex class II molecules were within cytoplasmic compartments consistent with a poor allostimulatory capacity . Bacteria or cytokines were very efficient in inducing progression from stage 1 towards stage 2 (mature) . Morphological changes were observed by confocal analysis including depolymerization of F-actin and loss of vinculin containing adhesive structures which correlates with acquisition of high motility . Antigen uptake and presentation of native protein antigen was reduced . In contrast, presentation of immunogenic peptides and allostimulatory activity became very efficient and secretion of IL-12 p75 was detectable after antigen presentation . This functional DC maturation ended by apoptotic cell death, and no reversion to the immature phenotype was observed. Gene, 1997 Jan 15, 184(2), 291 - 8 Physical and genetic map of the chromosome of Dichelobacter nodosus strain A198; La Fontaine S et al.; A physical map of the chromosome of Dichelobacter nodosus strain A198 was constructed using the restriction endonucleases EagI and StuI . Mapping data indicated the presence of a single, circular chromosome of 1.54 Mb . The three rRNA operons and the virulence related locus (vrl) were precisely positioned at the junctions of EagI and StuI fragments, and their transcriptional orientations were also determined . Other D . nodosus genes were assigned to specific EagI and StuI fragments . Analysis of the resultant map revealed that the putative virulence genes were not clustered on the chromosome which suggests that the D . nodosus virulence determinants have been acquired gradually and that virulence in D . nodosus is an evolving trait. Eur J Biochem, 1997 Jan 15, 243(1-2), 474 - 81 Assignment of the ligand geometry and redox potentials of the trihaem ferricytochrome c3 from Desulfuromonas acetoxidans; Turner DL et al.; Cytochrome c551.5 is a trihaem cytochrome of the cytochrome c3 family isolated from Desulfuromonas acetoxidans . Although several X-ray structures are available for tetrahaem cytochromes of this family, there is no X-ray structure for trihaem cytochromes . Cytochrome C551.5 was studied in the oxidized form by means of two-dimensional NMR . The pattern of observed interhaem NOESY connectivities is in agreement with the haem core structure previously determined by NMR for the reduced protein {Coutinho, I . B., Turner, D . L., Liu, M . Y., LeGall, J . & Xavier, A . V . (1996) J . Biol . Inorg . Chem . 1, 305-311} . The similarities found between the haem core structure and the amino acid sequence of cytochrome c551.5 and those of tetrahaem cytochromes c3 allows each of the haems to be specifically assigned in the polypeptide sequence, and the attribution of the midpoint redox potentials to the individual haems . This also allows individual redox potentials to be assigned to each haem in the NMR spectrum . The paramagnetic shifts of the 13C resonances of the haem substituents were analyzed in terms of pi molecular orbitals with perturbed D4h symmetry . The parameters of this analysis have been shown to be controlled by the orientation of the axial ligands in several other bis-His-coordinated haems and hence the ligand geometry was deduced for cytochrome C551.5 . The structural analogy between the relative haem plane orientations in cytochrome c551.5 and the tetrahaem cytochromes c3 is found to extend to the axial ligands with the largest differences being in the vicinity of the deleted fourth haem, using the numbering of cytochrome c3 haems. Eur J Biochem, 1997 Jan 15, 243(1-2), 393 - 9 An ultracentrifugal approach to quantitative characterization of the molecular assembly of a physiological electron-transfer complex: the interaction of electron-transferring flavoprotein with trimethylamine dehydrogenase; Wilson EK et al.; The interaction between two physiological redox partners, trimethylamine dehydrogenase and electron-transferring flavoprotein, has been characterized quantitatively by analytical ultracentrifugation at 4 degrees C . Analysis of sedimentation-equilibrium distributions obtained at 15 000 rpm for mixtures in 10 mM potassium phosphate, pH 7.5, by means of the psi function {Wills, P . R., Jacobsen, M . P . & Winzor, D . J . (1996) Biopolymers 38, 119-130} has yielded an intrinsic dissociation constant of 3-7 microM for the interaction of electron-transferring flavoprotein with two equivalent and independent sites on the homodimeric enzyme . This investigation indicates the potential of sedimentation equilibrium for the quantitative characterization of interactions between dissimilar macromolecules. Eur J Biochem, 1997 Jan 15, 243(1-2), 209 - 18 Identification and structure of activated-platelet protein-1, a protein with RNA-binding domain motifs that is expressed by activated platelets; Houng AK et al.; Be