Mol Microbiol, 1994 May, 12(4), 613 - 9 Identification of lactoferrin-binding proteins from Treponema pallidum subspecies pallidum and Treponema denticola; Staggs TM et al.; Lactoferrin-binding or -associated proteins were identified in Treponema pallidum subspecies pallidum and Treponema denticola by affinity column chromatography using human lactoferrin and detergent-solubilized, radiolabelled spirochaetes . Two discrete polypeptides of T . pallidum with masses of 45 and 40 kDa and a broad band from 29-34 kDa exhibited association with human apo- and partially ferrated lactoferrin . T . denticola produced two proteins that associated with a lactoferrin affinity matrix (50 and 35 kDa) . T . pallidum and T . denticola did not associate with soluble, human transferrin in parallel experiments . Soluble human lactoferrin competed with all lactoferrin-associated proteins from T . pallidum and T . denticola in competitive-binding assays . However, the T . denticola proteins dissociated from a lactoferrin-affinity matrix in the presence of differing concentrations of unlabelled, soluble lactoferrin competitor . Treatment with phospholipase D altered migration of the diffuse 29-34 kDa band of T . pallidum suggesting that the polypeptide was lipid-modified . Each of the lactoferrin-binding proteins from T . pallidum and T . denticola reacted with pooled rabbit syphilitic antisera . The lactoferrin-binding proteins of T . pallidum reacted with human sera from patients at all stages of syphilis . In addition, a monoclonal antibody generated against the 45 kDa polypeptide of T . pallidum crossreacted with the 29-34 kDa protein. Mol Microbiol, 1994 May, 12(4), 547 - 60 Regulation of a restriction and modification system via DNA inversion in Mycoplasma pulmonis; Dybvig K et al.; An invertible DNA element of 6.8 kb, designated the hsd1 locus, was identified in the chromosome of Mycoplasma pulmonis . Infection of host cells with mycoplasma virus P1 revealed that the organism's restriction and modification (R-M) properties are controlled by inversion of hsd1 . The nucleotide sequence of hsd1 revealed several genes, the predicted amino acids of which bear striking similarity to the subunits of the type I R-M enzymes previously found only in enteric bacteria. New Horiz, 1994 May, 2(2), 156 - 63 Is early feeding beneficial? How early is early? Minard G, Kudsk KA. Nutritional support is a vital part of the therapy of most hospitalized patients . Early initiation, particularly via the enteral route, has a significant effect on septic complications in a wide variety of patients . Early enteral feeding may also attenuate the hypercatabolic response that follows burn injury, although the evidence is somewhat conflicting . The mechanisms for the benefit of early nutrition have not been fully elucidated . However, preservation of gut mass, prevention of increased gut permeability to bacteria and other toxins, and maintenance of the gut-associated lymphoid tissue all probably play a role . The question "How early is early?" is important, since maintenance of early feeding requires time, patience, and dedication . It appears that starting nutrition within 24 hrs of major surgery, injury, or burn is ideal, but within 48 hours is acceptable . However, hemodynamic stability is a prerequisite to initiation of enteral feeding . Although labor intensive, the provision of early feeding, particularly via the enteral route, is a worthwhile goal for all clinicians. Curr Opin Nephrol Hypertens, 1994 May, 3(3), 356 - 63 Tubulointerstitial nephritis and vasculitis; Hill GS; This review discusses recent developments in infectious tubulointerstitial nephritis, including bacterial virulence factors, the interaction between leukocytes and bacteria, and early events in inflammation . In the area of vasculitis, little-studied specificities of antineutrophil cytoplasmic antibodies (ANCAs), including lactoferrin and elastase, plus newly described ANCAs with alpha-enolase specificity are discussed, along with newer studies on the pathogenesis of ANCA-mediated damage and animal models for ANCA-related vasculitis. J Hosp Infect, 1994 May, 27(1), 61 - 7 Enteroviruses, endoscopy and infection control: an applied study; Hanson PJ et al.; Decontamination methods for medical equipment are based largely on bacterial studies yet enteroviruses are more resistant to disinfection than most vegetative bacteria and other viruses . To study the elimination of enteroviruses from endoscopes, poliovirus was aspirated into the suction-biopsy channels of five gastroscopes . Endoscopes were cleaned in detergent and disinfected in 2% alkaline glutaraldehyde . Contamination was measured before and after cleaning and after various periods of disinfection by irrigating the channels with viral medium and quantifying surviving virus by plaque assay . The effectiveness of glutaraldehyde against cell-free and cell-associated polio virus, dried to a surface in a protein coagulum, was also studied . Cleaning reduced virus by a mean of 4.6 log10 plaque forming units (pfu) ml-1 . Samples were virus-free after 2 min disinfection . Virus dried on surfaces was inactivated in 1 min by 2% and 1% glutaraldehyde, with a reduction of > 6 log10 pfu ml-1 . Thus, cleaning was effective against heavy viral contamination while glutaraldehyde rapidly inactivated poliovirus even when dried to a surface in serum. Biochem Mol Biol Int, 1994 May, 33(1), 45 - 54 Parallel up-regulation of CD11B and CD45 on neutrophilic leukocytes exposed to soluble factors of oral pathogens; Cui Y et al.; We tested culture supernatants from a battery of oral bacterial strains for their ability to influence the expression of CD11b and CD45 on the neutrophil plasma membrane . Several bacterial extracts stimulated the up-regulation of both CD11b and CD45 simultaneously . Two supernatants in particular (a clinical isolate of A . actinomycetemcomitans and F . nucleatum ATCC25586) potently stimulated the deployment of CD11b and CD45 from their intracellular storage site to the plasma membrane . Both supernatants inhibited superoxide release stimulated by exposure of neutrophils to formyl methionyl leucyl phenylalanine (FMLP) but had variable effects on superoxide release stimulated by phorbol myristate acetate (PMA) . The ability of products of oral bacteria to modulate neutrophil plasma membrane antigen composition may regulate functional reactivity and thus be an important factor in the pathogenesis of periodontal infection and inflammation. Rev Inst Med Trop Sao Paulo, 1994 May-Jun, 36(3), 255 - 64 Etiology and severity of community acquired pneumonia in children from Uruguay: a 4-year study; Hortal M et al.; The 4-year study (1987-1990) covered the major clinical-epidemiological characteristics of pneumonia in children as diagnosed at the emergency service of the Children's Hospital, as well as etiologies, and factors involved in the most severe cases . Etiology was determined in 47.7% of the 541 pneumonia cases, involving 283 pathogens of which 38.6% were viruses and 12.6% bacteria . Viral and mixed etiologies were more frequent in children under 12 months of age . Bacteria predominated in ages between 6 and 23 months . Among the viruses, respiratory syncytial virus predominated (66%) . The bacterial pneumonias accounted for 12.2% of the recognized etiologies . The most important bacterial agents were S . pneumoniae (64%) and H . influenzae (19%) . H . influenzae and mixed infections had a relevant participation during the 1988 season, pointing to annual variations in the relative participation of pathogens and its possible implication in severity of diseases . Correlation of severity and increased percentage of etiological diagnosis was assessed: patients with respiratory rates over 70 rpm, or pleural effusion and/or extensive pulmonary parenchyma compromise yielded higher positive laboratory results . Various individual and family risk factors were recognized when comparing pneumonia children with healthy controls. J Biol Chem, 1994 Apr 29, 269(17), 12858 - 64 Antibodies against the C2 COOH-terminal region discriminate the active and latent forms of the multicatalytic proteinase complex; Arribas J et al.; The mouse cDNA homologues of the rat C2, C9, and C5 subunits of the multicatalytic proteinase have been cloned and expressed in bacteria . The respective recombinant proteins were purified and used to produce specific anti-subunit antibodies . Immunoblotting of two-dimensional gels of purified rat liver multicatalytic proteinase showed that the C2 (32-kDa) and C9 (29-kDa) polypeptides are resolved into three and two isoelectric variants, respectively, likely due to post-translational modifications, i.e . phosphorylation, and the presence of two anti-C5 reacting polypeptides (25.5 and 23 kDa) . Epitope mapping of the anti-C2-specific antibody with different constructs of the recombinant C2 protein allowed us to determine that one major epitope of this anti-C2 antibody is located within the last 9-11 amino acids of the C2 polypeptide . Affinity purified antibodies directed against the C2 COOH-terminal were able to discriminate the active and latent forms of the multicatalytic proteinase, supporting the conclusion that the C2 protein found in the active form of the enzyme is a polypeptide of 28 kDa, produced by the loss, at least, of the last 9-13 amino acids (DEPAEKADEPMEH) of the intact C2 (32-kDa) component . By in vitro treatment of the latent form of the enzyme with elastase, we show the conversion of the C2 (32-kDa) component to a 28-kDa protein with loss of recognition by the anti-C2 COOH-terminal affinity purified antibodies, but this limited degradation of the C2 component did not have any significant effect on the proteolytic activity (assayed with myelin basic protein and fluorogenic peptides) of the multicatalytic proteinase . It is suggested that the proteolytic cleavage of the C2 COOH-terminal region may be involved in the regulation of the interaction of the multicatalytic proteinase with other cellular proteins and/or in the translocation of the complex to the nucleus. Cell, 1994 Apr 22, 77(2), 195 - 205 An adenosine nucleotide switch controlling the activity of a cell type-specific transcription factor in B . subtilis; Alper S et al.; The sigma F factor establishes cell type-specific gene transcription during sporulation in B . subtilis . sigma F is negatively regulated by SpollAB, which forms complexes with sigma F or SpollAA . ATP and its nonhydrolyzable analogs stimulate the formation of the SpollAB.sigma F complex, whereas ADP stimulates the formation of the SpollAB.SpollAA complex . Which protein SpollAB associates with is determined by the concentrations of the two nucleotides, on which basis we propose a partner-switching model for the regulation of sigma F: {formula: see text} Consistent with this model, SpollAA reverses SpollAB-mediated inhibition of sigma F-directed transcription in a manner that depends on ADP . Cell-specific activation of sigma F could be due to an alteration in adenosine nucleotide levels in one cell of the sporangium. Biochim Biophys Acta, 1994 Apr 20, 1191(1), 43 - 51 Liposome-complement interactions in rat serum: implications for liposome survival studies; Devine DV et al.; Serum complement opsonizes particles such as bacteria for clearance by the reticuloendothelial system . Complement has been reported to interact with liposomes and therefore may mediate the reticuloendothelial system clearance of liposomes . This study has used a rat serum model to define some of the characteristics of liposomes which modulate their ability to activate complement . Using functional hemolytic assays and C3/C3b crossed immunoelectrophoresis, we have demonstrated that liposomes activated rat complement in a dose-dependent manner with higher concentrations of liposomes activating higher levels of complement . The detection of complement activation required the inclusion of phospholipids bearing a net charge . Complement activation occurred via the classical pathway; no alternative pathway activation was detected . The presence of cholesterol contributed to complement activation in a dose-dependent manner . Phospholipid fatty acyl chain length did not influence complement activation while the introduction of unsaturated acyl chains markedly decreased levels of complement activation . Liposome size also influenced complement activation with 400 nm unilamellar vesicles more effectively activating complement than 50 nm vesicles for equivalent amounts of exposed lipid . These studies demonstrate that the composition of the liposome greatly affects the in vitro activation of rat serum complement and suggest that the biological half-life of liposomes in the circulation of rats may be altered by changing the liposome composition to reduce complement activation. Biochemistry, 1994 Apr 19, 33(15), 4714 - 20 Mechanism for the desulfurization of L-cysteine catalyzed by the nifS gene product; Zheng L et al.; The nifS gene product (NIFS) is a pyridoxal phosphate binding enzyme that catalyzes the desulfurization of L-cysteine to yield L-alanine and sulfur . In Azotobacter vinelandii this activity is required for the full activation of the nitrogenase component proteins . Because the nitrogenase component proteins, Fe protein and MoFe protein, both contain metalloclusters which are required for their respective activities, it is suggested that NIFS participates in the biosynthesis of the nitrogenase metalloclusters by providing the inorganic sulfur required for Fe-S core formation {Zheng, L., White, R . H., Cash, V . L . Jack, R . F., & Dean, D . R . (1993) Proc . Natl . Acad . Sci . U.S.A . 90, 2754-2758} . In the present study the mechanism for the desulfurization of L-cysteine catalyzed by NIFS was determined in the following ways . First, the substrate analogs, L-allylglycine and vinylglycine, were shown to irreversibly inactivate NIFS by formation of a gamma-methylcystathionyl or cystathionyl residue, respectively, through nucleophilic attack by an active site cysteinyl residue on the corresponding analog-pyridoxal phosphate adduct . Second, this reactive cysteinyl residue, which is required for L-cysteine desulfurization activity, was identified as Cys325 by the specific alkylation of that residue and by site-directed mutagenesis experiments . Third, the formation of an enzyme-bound cysteinyl persulfide was identified as an intermediate in the NIFS-catalyzed reaction . Fourth, evidence was obtained for an enamine intermediate in the formation of L-alanine.(ABSTRACT TRUNCATED AT 250 WORDS) Biochem Biophys Res Commun, 1994 Apr 15, 200(1), 435 - 42 Yge1p, a eukaryotic Grp-E homolog, is localized in the mitochondrial matrix and interacts with mitochondrial Hsp70; Nakai M et al.; Yeast Yge1p, the gene product of YGE1, is a eukaryotic GrpE homolog found recently (Ikeda et al., 1994) . We have revealed here that Yge1p is a soluble protein in the mitochondrial matrix . Depletion of Yge1p in yeast cells resulted in accumulation of the precursor of F1-ATPase beta subunit in vivo, suggesting that Yge1p is involved in protein import into mitochondria . Overexpression of Yge1p in the temperature-sensitive mutant strains of mitochondrial hsp70, Ssc1p, caused hypersensitivity to temperature for cell growth, suggesting a genetic interaction between the YGE1 and SSC1 genes . A physical interaction between Yge1p and Ssc1p was directly demonstrated by co-immunoprecipitation of Ssc1p by the anti-Yge1p antibodies . We propose that Yge1p functions in cooperation with Ssc1p in a similar manner as bacterial GrpE with DnaK. J Mol Biol, 1994 Apr 15, 237(5), 622 - 40 Directed mutational analysis of bacteriochlorophyll a biosynthesis in Rhodobacter capsulatus; Bollivar DW et al.; Previous studies have established that most if not all of the genes required for synthesis of the Rhodobacter capsulatus essential photosystem are clustered on a 46 kb region of the chromosome known as the photosynthesis gene cluster . This region has recently been sequenced in its entirety by Hearst and co-workers, revealing the existence of 23 open reading frames, many of which are thought to be involved in the synthesis of bacteriochlorophyll . In this study we have undertaken a systematic directed mutational analysis of 12 open reading frames in the photosynthesis gene cluster to evaluate whether individual open reading frames have a role in photopigment biosynthesis . The results of this analysis demonstrate that mutations constructed in seven open reading frames resulted in a loss of bacteriochlorophyll biosynthesis, concomitant with the accumulation of specific intermediates in the Mg-tetrapyrrole biosynthetic pathway . One mutation was observed to result in partial disruption of bacteriochlorophyll biosynthesis, leading to the accumulation of bacteriochlorophyll as well as an intermediate in the biosynthetic pathway . We also observed that disruptions constructed in four open reading frames had no discernible effect on the synthesis of photopigments . The results of this analysis are discussed with regard to our current understanding of the role of each of these open reading frames in the synthesis of the R . capsulatus photosystem. J Biol Chem, 1994 Apr 15, 269(15), 11081 - 9 A reverse gyrase with an unusual structure . A type I DNA topoisomerase from the hyperthermophile Methanopyrus kandleri is a two-subunit protein; Kozyavkin SA et al.; Reverse gyrase, an ATP-dependent topoisomerase that positively supercoils DNA, has been purified to near-homogeneity from the hyperthermophile Methanopyrus kandleri . It migrates on SDS-polyacrylamide gel electrophoresis as two principal bands with apparent molecular masses of 150 and 50 kDa . Both proteins remain associated throughout all chromatographic steps . Transfer of a radioactive phosphate from DNA to the 50-kDa protein and gel retardation experiments indicate that this protein forms the covalent complex with DNA . A blot overlay assay identifies the 150-kDa protein as the potential ATPase . This is the first evidence that a reverse gyrase can be a topoisomerase consisting of two protomers . In analogy with the DNA gyrase A subunit (DNA breakage and reunion activity) and the B subunit (ATPase), the 50- and 150-kDa components of Mka reverse gyrase have been designated the A and B subunits, respectively . Methanopyrus reverse gyrase changes DNA linking number in steps of one and its A subunit covalently binds to the 5'-DNA phosphoryl group . It nicks DNA at sites that predominantly have a cytosine at the -4-position . The same rule was derived previously for monomeric reverse gyrase from sulfur-metabolizing hyperthermophiles and for topoisomerase I from mesophilic bacteria . Based on these results, Mka reverse gyrase is classified as belonging to group A of type I topoisomerases . The structural diversity of type I group A topoisomerases parallels the diversity of type II enzymes and suggests the evolution of an essential function by gene fusion. Am J Ophthalmol, 1994 Apr 15, 117(4), 480 - 7 Vaccinia keratouveitis manifesting as a masquerade syndrome; Lee SF et al.; A patient who used contact lenses and had a history of blunt trauma developed vaccinia keratouveitis after accidental ocular autoinoculation from a recent vaccination site . Corneal and conjunctival cultures were taken for bacteria, fungi, Acanthamoeba, and viruses . Viral-like cytopathic effects became evident in tissue culture within three days . Immunofluorescence studies were negative for varicella-zoster virus, herpes simplex virus, adenovirus, measles, mumps, parainfluenza, and influenza . Pox viral particles were identified in the infected tissue cultures by electron microscopy . The Hind III restriction endonuclease profile of the viral DNA isolate was similar to the Lister strain of vaccinia virus . Ocular vaccinia may manifest as a masquerade syndrome and may mimic signs of herpes simplex virus, varicella-zoster virus, and Acanthamoeba infection . Although vaccination with vaccinia is currently limited to a few populations throughout the world, vaccinia must still be considered in the differential diagnosis of infectious keratouveitis. Neuroreport, 1994 Apr 14, 5(8), 977 - 80 Inducible microglial nitric oxide synthase: a large membrane pool; Wood PL et al.; Microglia are the only immunocompetent cells resident in the central nervous system which are capable of protecting the brain from infection and tumors . These resident macrophages possess a vast array of mechanisms for the destruction of bacteria and tumor cells . One of these mechanisms involves the generation of nitric oxide which can kill cells by inhibition of glycolysis, the TCA cycle and DNA synthesis . In this regard, we demonstrate, for the first time, that the inducible form of nitric oxide synthase (NOS) in microglia involves both cytosolic and membrane bound pools . Both pools of NOS were potently and stereo-specifically inhibited by NOS inhibitors . In addition, while these pools were unaffected by Ca2+, they were partially inhibited by calmodulin antagonists . These data would suggest that inducible NOS in lipopolysaccharide (LPS) treated microglia, constitutes two major compartments and may involve a novel isoform which is membrane associated . With regard to the possible physiological relevance for the membrane-bound NOS, we speculate that this presents an efficient means of supplying nitric oxide to the extracellular environment where it could gain rapid access to tumors and bacteria . This would result in inhibition of cellular function in these invading cells while limiting access of nitric oxide to the intracellular environment of microglia where NO could lead to depressed microglial function. Proc Natl Acad Sci U S A, 1994 Apr 12, 91(8), 2965 - 9 Ankyrin binds to two distinct cytoplasmic domains of Na,K-ATPase alpha subunit; Devarajan P et al.; Ankyrin has emerged as a ubiquitous protein linking integral membrane transport proteins such as Na,K-ATPase to an underlying spectrin cytoskeleton . This interaction is mediated by the alpha subunit of Na,K-ATPase; however, the nature of the ankyrin binding site in Na,K-ATPase is unknown . As a step to determine the mechanism of this interaction, the ankyrin binding region of human erythrocyte spectrin and each of five putative cytoplasmic domains of the Na,K-ATPase alpha subunit have been prepared as recombinant fusion proteins in bacteria and analyzed for their interaction with erythrocyte and kidney ankyrin (Ank1 and Ank3, respectively) in vitro . Spectrin binds both Ank1 and Ank3 avidly, as expected . Two of the Na,K-ATPase domains, immobilized on a bioaffinity column, also interact specifically with both of these ankyrins . These ATPase domains are encoded by codons 140-290 (domain II) and 345-784 (domain III), with domain II displaying the greatest apparent affinity . Sequences in domain II are highly conserved between species and isoforms of Na,K-ATPase and are homologous to a cytoplasmic domain in H,K-ATPase and to a limited region of sequence in Ca-ATPase . Conversely, domain II shares no significant homology with other ankyrin binding proteins such as band 3 and Na(+)-channel proteins . These results identify a clear function for a conserved but previously not understood region of the alpha subunit of Na,K-ATPase and suggest that the interaction of ankyrin with membrane transport proteins may involve complex tertiary structural determinants not easily deduced from the primary sequence. Biochim Biophys Acta, 1994 Apr 6, 1217(3), 257 - 65 Molecular cloning and characterization of an isoprenylated 67 kDa protein; Asundi VK et al.; The cDNA coding for a 67 kDa protein (p67) was isolated from a rat Schwann cell library . A recombinant form of p67 expressed in bacteria was used to produce polyclonal anti-p67 antibodies . By immunoblot analysis p67 was found to be expressed in most tissues and cell lines examined . Inspection of the deduced amino acid sequence revealed a COOH-terminal consensus sequence for isoprenylation . Consistent with this finding, p67 was a substrate for isoprenylation in vitro by geranylgeranylpyrophosphate . p67 was associated predominantly with the particulate fraction of rat smooth muscle cells . The rat p67 sequence was highly homologous to a family of recently described human and mouse gamma-interferon inducible, guanine nucleotide binding proteins. Eur J Immunogenet, 1994 Apr, 21(2), 125 - 31 Dual role of mannan-binding protein in infections: another case of heterosis? Garred P, Harboe M, Oettinger T, Koch C, Svejgaard A. Human mannan-binding protein (MBP) is a serum lectin that participates in the immune defence by mediating phagocytosis and activation of complement . Variant MBP alleles causing dominant low-serum concentrations have high frequencies in all populations studied, and therefore, low MBP concentrations may confer selective advantages to those individuals carrying the variant alleles . Mycobacterium leprae, the causative agent of leprosy, is an obligate intracellular parasite dependent on phagocytosis to invade host cells . The serum concentrations of MBP in 36 Ethiopian patients (median: 1688 micrograms l-1) with lepromatous or borderline lepromatous leprosy were significantly (P < 0.001) higher than in 26 healthy Ethiopian blood donors (median: 368 micrograms l-1) . Only 17% of the patients vs . 58% of the donors (P = 0.0019) had the relatively low MBP concentrations usually associated with variant alleles . Functional studies revealed that M . leprae and M . tuberculosis sonicates bind MBP as strongly as pure mannan . These observations suggest a role for mycobacteria as a selective force in the positive selection of alleles causing low levels of MBP and warrant genetic studies of patients infected with these bacteria. Infect Control Hosp Epidemiol, 1994 Apr, 15(4 Pt 1), 253 - 64 Risk factors for central venous catheter-related infections in surgical and intensive care units . The Central Venous Catheter-Related Infections Study Group; Moro ML et al.; OBJECTIVE: To identify avoidable risk factors for central venous catheter (CVC) infections in patients undergoing short-term catheterization . DESIGN: Prospective multicenter cohort study . SETTING: Two university teaching hospitals and five large nonteaching hospitals . PATIENTS: Patients admitted to intensive care units or surgical units and exposed to short-term CVCs . RESULTS: Of 623 catheterization episodes, 9.3% were associated with catheter-related infections (CRI) . The skin at the catheter site was frequently colonized (16.2%) and was the potential source of infection in 56.1% of the cases, mostly local infections . The hub was colonized less frequently (3.5%) but was responsible for systemic infections more frequently . The following variables were independently associated with CRI: duration of catheterization (for 7 to 14 days, odds ratio {OR}, 3.9; 95% confidence interval {CI}95, 1.4 to 10.7; and for > 14 days, OR, 5.1; CI95, 1.7 to 15.4), coronary care unit service (OR, 6.7; CI95, 1.1 to 42.9) or surgery service (OR, 4.4; CI95, 1.03 to 18.5), second episode of catheterization (OR, 7.6; CI95, 1.8 to 32.3), skin colonization at the insertion site (OR, 56.5; CI95, 10.8 to 296), and hub colonization (OR, 17.9; CI95, 2.4 to 132) . The risk associated with skin colonization varied with use of jugular access or simultaneous colonization of the hub . When only symptomatic CRI was considered, the risk associated with hub colonization was consistently higher (OR, 36.6; CI95, 7 to 190) than that associated with skin colonization (OR, 3.2; CI95, 0.7 to 14) . Age, transparent dressing, jugular insertion, male gender, duration of catheterization, and hub colonization were independent risk factors for skin colonization . The effect of age varied by type of dressing and vice versa; the effect of jugular access varied by sex; and the effect of transparent dressing varied by length of catheterization and vice versa . Total parenteral nutrition and skin colonization were independently associated with an increased risk of hub colonization . CONCLUSIONS: Skin and hub colonization are the two major determinants for endemic CRIs; colonization of the hub, however, is more frequently associated with more severe infections . In order to reduce CRIs, more efforts should be focused on understanding which factors increase the risk of colonization both of the skin and of the hub. Nippon Rinsho, 1994 Apr, 52(4), 1110 - 7 {Pathogenesis of Graves' disease}; Yokoyama N et al.; The discovery of long acting thyroid stimulator in Graves' disease and autoantibodies specific for the thyroid in Hashimoto disease in 1956, were the earliest examples of autoimmune responses . Autoimmune thyroid disease has many important advantages in the investigation of autoimmune disease when compared to the other disease . It is possible to obtain thyroid tissue at biopsy and to investigate the histology by various methods and the interactions between thyrocytes and infiltrated mononuclear cells in vitro . Important autoantigens, such as the TSH receptor, thyroid peroxidase and thyroglobulin have already been cloned and each autoantigen has a specific function . Furthermore, we can observe precisely the clinical course of the disease using laboratory parameters . In this review, the pathogenesis of Graves' disease will be overviewed using the results obtained, mainly in our laboratory, in the following topics: (1) Immunogenetics: HLA class I and II, Gm, multiple genes (2) Trigger: bacteria, retrovirus (HIV, HTLV-I), radiation (3) Initiation and perpetuation of autoimmune responses: role of HLA class I and II antigens, characteristics of infiltrated mononuclear cells, interactions among thyrocytes, mononuclear cells and endothelial cells, role of cytokines, adhesion molecules (4) Autoantibodies: methods of determination and clinical correlates of TSH receptor antibodies (5) Autoantigens: structure and functional relationship of TSH receptor (6) Future studies: possible methods of treatment based on pathogenesis, a model of new treatment. J Clin Periodontol, 1994 Apr, 21(4), 240 - 9 Clinical and histological evaluation of ligature-induced periodontal breakdown in domestic ferrets immunosuppressed by Cyclosporin-A; Fischer RG et al.; The aim of this study was to evaluate, clinically and histologically, the effect of Cyclosporin-A (CyA) on the progression of the periodontal breakdown in the domestic ferret, using the ligature induced periodontitis model . At the start of the experiment (day 0), silk ligatures were placed at the gingival margin level of experimental teeth . The contralateral teeth served as non-ligated control teeth . Clinical measurements included gingival index, probing pocket depth (PPD), probing attachment level (PAL) and gingival overgrowth (GO) and they were performed on days 0, 14 and 28 . Cyclosporin-A, 10 mg/kg/d, was given subcutaneously from day 0 to 21, while in the last week there was a reduction of this dose to 5 mg/kg/d . Blood samples were taken on days 0, 14 and 28 . On day 28 the animals were sacrificed . Histological sections were prepared for light microscopy . The histometric measurements performed were: (1) the distance between cemento-enamel junction and the alveolar bone crest and (2) loss of connective tissue attachment . The number of sections with root resorption areas was observed . Cell counts were taken in 4 different areas . The results showed, on days 14 and 28, a significant increase of the mean values of PPD and PAL at experimental teeth as compared to the control teeth . GO was present on experimental teeth on days 14 and 28 . On day 28, the control teeth presented a very small increase in GO, mainly at the buccal sites of P4 and M1 . The histometric results showed a significant loss of attachment and bone resorption in the experimental teeth . Root resorption was found in experimental teeth only . The predominant inflammatory cell in the 4 experimental areas was polymorphonuclears.(ABSTRACT TRUNCATED AT 250 WORDS) Int J Syst Bacteriol, 1994 Apr, 44(2), 308 - 14 Phylogenetic evidence for Sphingomonas and Rhizomonas as nonphotosynthetic members of the alpha-4 subclass of the Proteobacteria; Takeuchi M et al.; To clarify the taxonomic relationships of the genera Rhizomonas and Sphingomonas, the 16S rRNA sequence of Rhizomonas suberifaciens IFO 15211T (T = type strain) was determined . A phylogenetic analysis of aligned 16S rRNA gene sequences revealed that eight species of the genus Sphingomonas and R . suberifaciens are closely related to Erythrobacter longus and Porphyrobacter neustonensis and, therefore, belong in the alpha-4 subclass of the Proteobacteria . Within this subclass, Sphingomonas species and R . suberifaciens are phylogenetically interrelated and comprise several subgroups . Our findings show that the genus and species definitions of these organisms are in need of revision. FEMS Microbiol Lett, 1994 Apr 1, 117(2), 157 - 61 Some rumen ciliates have endosymbiotic methanogens; Finlay BJ et al.; Most of the small ciliate protozoa, including Dasytricha ruminantium and Entodinium spp . living in the rumen of sheep, were found to have intracellular bacteria . These bacteria were not present in digestive vacuoles . They showed characteristic coenzyme F420 autofluorescence and they were detected with a rhodamine-labelled Archaea-specific oligonucleotide probe . The measured volume percent of autofluorescing bacteria (1%) was close to the total volume of intracellular bacteria estimated from TEM stereology . Thus it is likely that all of the bacteria living in the cytoplasm of these ciliates were endosymbiotic methanogens, using H2 evolved by the host ciliate to form methane . Intracellular methanogens appear to be much more numerous than those attached to the external cell surface of ciliates. Am J Physiol, 1994 Apr, 266(4 Pt 1), C877 - 92 Macromolecular crowding and confinement in cells exposed to hypertonicity; Garner MM et al.; The nonideal properties of solutions containing high concentrations of macromolecules can result in enormous increases in the activity of the individual macromolecules . It has been proposed that molecular crowding and confinement occur in cells and are major determinants of the activity of the proteins and other intracellular macromolecules . This concept has important implications for cell volume regulation because, under crowded conditions, relatively small changes in concentration, consequent to alterations of water content, lead to large changes in macromolecular activity . This review considers several aspects of macromolecular crowding and confinement, including: 1) the physical chemical principles involved; 2) in vitro demonstrations of the effects; 3) relation to water activity; 4) estimates of the actual intracellular activity of water and macromolecules; 5) relation to osmotic regulation in various types of cells, including bacteria, red blood cells, and complex nucleated cells; and 6) the relation to inorganic ions and organic osmolytes in cells stressed by hypertonicity . We conclude that, while there is compelling evidence for important effects of molecular crowding in vitro and in red blood cells, the role of macromolecular crowding and confinement in osmotic regulation of more complex cells is an open question that deserves the extensive attention it is currently receiving. Lab Invest, 1994 Apr, 70(4), 572 - 8 Idiotype identity in a MALT-type lymphoma and B cells in Helicobacter pylori associated chronic gastritis; Greiner A et al.; BACKGROUND: To investigate the mechanisms triggering MALT-lymphoma development, we examined the occurrence of normal B cells in lymphoid tissue and chronic gastritis with the same idiotype as an IgA-positive MALT lymphoma . EXPERIMENTAL DESIGN: Lymphoma idiotype IgA was produced by monoclonal human antibody technology . Against this idiotype a murine monoclonal antibody 27/165 with anti-idiotypic (alpha Id) specificities was raised, and applied immunohistochemically to identify the non-neoplastic precursor B cells in non-neoplastic human tissues . RESULTS: alpha Id 27/165 reacted exclusively with the IgA expressing MALT lymphoma but not with 20 other MALT-type gastric lymphomas nor with 26 nodal lymphomas and was not reactive with normal and inflamed lymph nodes . alpha Id 27/165 immunoreactivity was also absent from MALT of different mucosal sites but was readily encountered on a substantial number of lymphocytes and plasma cells in 95% cases of chronic gastritis associated with Helicobacter pylori (H.p.) . The target antigen of the lymphoma IgA was found to be a common antigen of IgA and IgM plasma cells of MALT but not a constituent of bacteria commonly involved in the pathogenesis of gastritis . CONCLUSIONS: The distinct binding of alpha Id 27/165 to only reactive mucosal B cells is a first direct evidence for the evolution of MALT-type lymphoma from chronic gastritis . Since the target antigen of the lymphoma IgA has been found to be an autoantigen of MALT plasma cells it is suggested that this MALT-type lymphoma may have arisen after triggering by an autoimmune response resulting from H.p.-induced gastritis. Ann Surg, 1994 Apr, 219(4), 389 - 99 Autoregulation by eicosanoids of human Kupffer cell secretory products . A study of interleukin-1, interleukin-6, tumor necrosis factor-alpha, transforming growth factor-beta, and nitric oxide; Roland CR et al.; OBJECTIVE: Methods employed previously to analyze the secretory behavior of rodent Kupffer cells (KC) were used to examine the human KC's secretory response to lipopolysaccharide (LPS) . SUMMARY BACKGROUND DATA: As the resident hepatic macrophage, the KC resides at the interface between the portal and systemic circulations . Consequently, this cell may play an integral role in the immune response to antigens and bacteria in the sinusoid . Study of cytokine production by the KC has relied predominantly on the rat as the source of these cells . Whether human KCs respond similarly to rat KCs after LPS stimulation has been a matter of speculation . METHODS: Kupffer cells obtained from seven human livers were tested under conditions identical to those used to study rat KCs . Kupffer cells rested for 12 hours after isolation were stimulated with LPS (2.5 micrograms/mL) . Arginine concentration in the culture medium varied from 0.01 to 1.2 mM . To examine the role of eicosanoids, parallel culture wells received indomethacin (10 microM) . Culture supernatants were assayed for interleukin-1 (IL-1), interleukin-6 (IL-6), tumor necrosis factor-alpha (TNF-alpha), transforming growth factor-beta (TGF-beta), prostaglandin E2 (PGE2), and nitric oxide . RESULTS: Similar to the rat KC, LPS-stimulated human KCs released IL-1, IL-6, TNF-alpha, TGF-beta, and PGE2 . However, unlike rat KCs, nitric oxide could not be detected, regardless of whether the human KCs were exposed to LPS, interferon-gamma (INF-gamma), or LPS + IFN-gamma . Similar to rat KCs, indomethacin prevented PGE2 release while significantly upregulating TNF-alpha, IL-1, and IL-6, but not TGF-beta, consistent with an autoregulatory control of eicosanoids over proinflammatory cytokines . As has been shown in the rat, physiologic levels of L-arginine (0.01 mM) significantly enhanced LPS-induced PGE2 secretion relative to the response in medium containing standard L-arginine concentration (1.2 mM); however, unlike the rat KC, the human's cytokine response to LPS was not downregulated by this enhanced PGE2 release . CONCLUSIONS: Although many functional features are shared by rat and human KCs, significant differences do exist . Such discrepancies reinforce the need to proceed with caution when generalizing from the results obtained in other species to human physiology. Dig Dis Sci, 1994 Apr, 39(4), 809 - 20 Long-term observations on morphological changes of choledochal epithelium after choledochoenterostomy in rats; Kurumado K et al.; Morphological changes of the common bile duct were observed macroscopically and microscopically 20 months after choledochojejunostomy and choledochocolonostomy in rats . The common bile ducts were remarkably dilated in all rats of both experimental models . Choledochal stones consisting of fatty acid calcium and calcium bilirubinate were seen in two of six rats with choledochojejunostomy and two of five rats with choledochocolonostomy . The main pathological change observed in both the groups was remarkable hyperplasia of the choledochal epithelium . Only one rat with choledochojejunostomy revealed normal epithelium with choledochal stone formation . Intestinal metaplasia was seen in two rats with choledochojejunostomy and all with choledochocolonostomy . Squamous pseudostratification of the epithelium indicating atypism was observed in two rats with choledochojejunostomy . Sialomucin producing cells and sulfomucin producing cells were seen in the hyperplastic portion of the epithelium . No malignant alteration of the epithelium was detected . These findings indicate that long-lasting exposure to digestive enzymes and bacteria causes epithelial hyperplasia and further exposure to digestive enzymes plays a major role in appearance of the epithelial atypism . Carcinogenesis of the choledochal epithelium under such an environment will need much more time to be established. J Bacteriol, 1994 Apr, 176(7), 1957 - 68 Genetics of the serine cycle in Methylobacterium extorquens AM1: identification of sgaA and mtdA and sequences of sgaA, hprA, and mtdA; Chistoserdova LV et al.; In a previous paper, we reported identification of the 5' part of hprA of Methylobacterium extorquens AM1, which encodes the serine cycle enzyme hydroxypyruvate reductase (L . V . Chistoserdova and M . E . Lidstrom, J . Bacteriol . 174:71-77, 1992) . Here we present the complete sequence of hprA and partial sequence of genes adjacent to hprA . Upstream of hprA, the 3' part of an open reading frame was discovered, separated from hprA by 263 bp . This open reading frame was identified as the gene encoding another serine cycle enzyme, serine glyoxylate aminotransferase (sgaA) . Cells containing an insertion mutation into sgaA were unable to grow on C1 compounds, demonstrating that the gene is required for C1 metabolism . Sequencing downstream of hprA has revealed the presence of another open reading frame (mtdA), which is probably cotranscribed with hprA . This open reading frame was identified as the gene required for the synthesis of 5,10-methylenetetrahydrofolate dehydrogenase . Our data suggest that this enzyme plays an integral role in methylotrophic metabolism in M . extorquens AM1, either in formaldehyde oxidation or as part of the serine cycle. Mol Cell Biol, 1994 Apr, 14(4), 2713 - 21 Regulation of cyclin D-dependent kinase 4 (cdk4) by cdk4-activating kinase; Kato JY et al.; The accumulation of assembled holoenzymes composed of regulatory D-type cyclins and their catalytic partner, cyclin-dependent kinase 4 (cdk4), is rate limiting for progression through the G1 phase of the cell cycle in mammalian fibroblasts . Both the synthesis and assembly of D-type cyclins and cdk4 depend upon serum stimulation, but even when both subunits are ectopically overproduced, they do not assemble into complexes in serum-deprived cells . When coexpressed from baculoviral vectors in intact Sf9 insect cells, cdk4 assembles with D-type cyclins to form active protein kinases . In contrast, recombinant D-type cyclin and cdk4 subunits produced in insect cells or in bacteria do not assemble as efficiently into functional holoenzymes when combined in vitro but can be activated in the presence of lysates obtained from proliferating mammalian cells . Assembly of cyclin D-cdk4 complexes in coinfected Sf9 cells facilitates phosphorylation of cdk4 on threonine 172 by a cdk-activating kinase (CAK) . Assembly can proceed in the absence of this modification, but cdk4 mutants which cannot be phosphorylated by CAK remain catalytically inactive . Therefore, formation of the cyclin D-cdk4 complex and phosphorylation of the bound catalytic subunit are independently regulated, and in addition to the requirement for CAK activity, serum stimulation is required to promote assembly of the complexes in mammalian cells. Cancer Res, 1994 Apr 1, 54(7 Suppl), 1948s - 1951s Experimental evidence for inhibition of N-nitroso compound formation as a factor in the negative correlation between vitamin C consumption and the incidence of certain cancers; Mirvish SS; Ascorbic acid (ASC) consumption is negatively correlated with the incidence of certain cancers . This is a review and update of the theory, which has recently been neglected, that this negative correlation is due to ASC inhibition of in vivo nitrosation . The review covers the older literature on ASC inhibition of carcinogenesis by nitrite administered with amines or amides and more recent studies on ASC inhibition of nitrosation by bacteria, nitrogen oxides, and activated macrophages; the role of oxygen in ASC inhibition of gastric nitrosation; ASC inhibition of N-nitrosoproline formation in subjects from areas with high incidences of certain cancers; dose and temporal relationships between ASC and in vivo nitrosation in humans; the role of substances other than ASC in the inhibition of nitrosation by vegetables and fruits; and the active secretion of ASC into the human stomach. Virology, 1994 Apr, 200(1), 313 - 8 The Epstein-Barr virus-associated protein p105 is not encoded by the Epstein-Barr virus genome; Turk SM et al.; Rabbit antibodies made to an Epstein-Barr virus (EBV)-associated hydrophobic protein p105 that cross-reacts antigenically with the herpes simplex virus glycoprotein gB inhibited the ability of EBV to induce immunoglobulin synthesis by normal B cells . Sequencing of p105 indicated that it was a keratin-like protein and not encoded by EBV . Analysis of EBV-producing cells with and without mycoplasma indicated that p105 is probably a mycoplasma protein that associates with the EBV virion. Minerva Anestesiol, 1994 Apr, 60(4), 157 - 64 {Evaluation of an experimental model of multiple organ dysfunction}; Di Filippo A et al.; OBJECTIVE . To perform an experimental model of Multiple Organ Dysfunction Syndrome (MODS) without employing bacteria or endotoxin stimulus and to follow its evolution in vivo by a Computerized Tomography analysis of the lungs . DESIGN . Rats were submitted to intraperitoneal injection of a 2.5% zymosan suspension in mineral oil (1 g/kg weight) or mineral oil alone; control rats received no treatment . METHODS . The observation period was 15 days . During this period symptoms and survival were noticed daily . CT scans of lungs were made at the 7th and 14th days; data were post-processed to obtain information on lung density . The rats were sacrificed at the 15th day by heart puncture; blood was utilized for determination of hemochrome, differential leukocyte count, thrombocytes, glycemia, uremia, bilirubin . Lungs, liver, spleen and kidney were dissected and weighted for determination of relative organ weight . DATA ANALYSIS . Data were compared by "t" Student's test for impaired data and Fisher Exact test . RESULTS . Symptoms, survival, blood analysis and relative organ weight agreed with a progressive, ingravescent, triphasic illness caused by a systemic inflammatory response involving remote organ too . The CT study proved able to monitoring and analyzing organ damage: a temporal sequence of evolution of damage exists; organ damage is localized in microcirculatory system (density augment) and in parenchyma (morphologic alterations and fibrosis) . DISCUSSION . The described experimental model reproduces a MODS-like illness in zymosan receiving rats; the CT scan is effective to evaluate the evolution of organ damage. Biochem Mol Biol Int, 1994 Apr, 32(6), 1085 - 92 Human leukemic pre-B line (KM-3) treated with phorbol-ester: trend of polyamines during cell differentiation; Trubiani O et al.; Aliphatic polyamines, putrescine, spermine and spermidine present in bacteria and eukaryotic cells are essential for cell growth . Generally, polyamine levels are elevated in rapidly growing normal and pathological systems . Since polyamines belong to the category of molecules whose synthesis is strongly activated during the G1 period they have been implicated in the cell's preparation for DNA replication . In our experiments, we have differentiated by phorbol-ester, the human pre-B leukemic cell line KM-3 . Biochemical and flow cytometric analysis show an increase, in treated cells, of polyamines pathway related to G1 and early S-phase of cell cycle during B cell differentiation. Hybridoma, 1994 Apr, 13(2), 123 - 30 Characterization of monoclonal antibodies to conserved antigens of Entamoeba histolytica and E . dispar and development of a stool ELISA; Sharma M et al.; Eight murine monoclonal antibodies (MAbs) were generated against Entamoeba histolytica NIH:200 . All MAbs reacted with three axenic strains of E . histolytica, NIH:200, HM-1: IMMS, and SAW 1734R clAR, but not with enteric bacteria or Giardia lamblia . Five of the MAbs reacted with low-molecular-weight, periodate-sensitive antigens of 14-21 kD, while one (AB31) reacted with high-molecular-weight, 90 to 200-kD protein determinants . MAb PC14 appeared to be specific for antigen in its native state . Another MAb (BB12) agglutinated live trophozoites and caused membrane fluorescence in contrast to the five other MAbs tested . Although BB12 reacted with the same 14 to 21-kD band on Western blot as AC55, the latter reacted with different cytoplasmic epitopes . All the MAbs reacted to five pathogenic (E . histolytica) and six nonpathogenic (E . dispar) clinical isolates . These MAbs may be helpful for studying conserved antigens of E . histolytica and were used to develop a sandwich ELISA for the diagnosis of intestinal amoebiasis . The assay was sensitive to 60 ng of E . histolytica antigen . A comparative study of microscopic examination of stool samples and the sandwich ELISA was conducted on 102 samples from patients with gastrointestinal complaints . The ELISA could detect all microscopically positive samples with a sensitivity of 100% and specificity of 93% . A sandwich ELISA using a monoclonal antibody to conserved antigens of E . histolytica has the potential to be a reliable method for the diagnosis of intestinal amoebiasis. Hum Gene Ther, 1994 Apr, 5(4), 469 - 80 An early history of gene transfer and therapy; Wolff JA et al.; The term "gene therapy" was coined to distinguish it from the Orwellian connotations of "human genetic engineering," which, in turn, was derived from the term "genetic engineering." Genetic engineering was first used at the Sixth International Congress of Genetics held in 1932 and was taken to mean "the application of genetic principles to animal and plant breeding." Once the basics of molecular genetics and gene transfer in bacteria were established in the 1960s, gene transfer into animals and humans using either viral vectors and/or genetically modified cultured cells became inevitable . Despite the early exposition of the concept of gene therapy, progress awaited the advent of recombinant DNA technology . The lack of trustworthy techniques did not stop many researchers from attempting to transfer genes into cells in culture, animals, and humans . Viral genomes were used for the development of the first relatively efficient methods for gene transfer into mammalian cells in culture . In the late 1970s, early transfection techniques were combined with selection systems for cultured cells and recombinant DNA technology . With the development of retroviral vectors in the early 1980s, the possibility of efficient gene transfer into mammalian cells for the purpose of gene therapy became widely accepted. Domest Anim Endocrinol, 1994 Apr, 11(2), 197 - 208 Absorption and metabolism of estrogens from the stomach and duodenum of pigs; Ruoff WL et al.; To determine the absorption and metabolism of 17 beta-estradiol (E2) by the stomach and liver of the pig, crystalline E2 was placed in the stomach of prepubertal gilts . Blood samples were subsequently obtained from the hepatic portal and jugular veins and plasma was assayed for E2, estrone (E1), 17 beta-estradiol-glucuronide (E2G), estrone-glucuronide (E1G) and estrone-sulfate (E1S) . Concentrations of E2, E1, E2G and E1S rose in the hepatic portal vein within five min and remained elevated for several hr . Concentration of E2 represented only 6% of the total estrogen detected in the hepatic portal vein during the sampling period, indicating that most of the E2 was converted or conjugated prior to entering the hepatic portal vein . The metabolism of E2 presumably occurred in the stomach mucosa because food had been withheld for 26 hr before infusion of E2 . Concentrations of E2G, E1G and E1S, but not E2 and E1, rose in the jugular vein and remained elevated for several hr . The lack of a rise in E2 and E1 in the jugular vein indicates that the E2 and E1 from the hepatic portal vein were completely converted and/or removed by the liver . Most of E2 was converted to E1 and then to E1G . The infusion of bile containing normal estrogens from pregnant gilts into the duodenum of prepubertal gilts resulted in a peak of E1G and E2G in the hepatic portal and jugular veins within a few minutes . This was followed in about 180 min by a second sustained rise . The first peak was essentially abolished by extracting E1 and E2 from the bile before infusion . The second peak failed to occur in gilts given antibiotics orally to reduce gut bacteria before infusion of bile. Vet Immunol Immunopathol, 1994 Apr, 40(4), 367 - 77 Uptake of ferritin and Bordetella avium in bronchus-associated lymphoid tissue of turkeys; Fagerland JA et al.; The uptake of macromolecular and particulate materials in bronchus-associated lymphoid tissue (BALT) in turkeys was examined using transmission electron microscopy . Tracer materials used were live and ultraviolet-killed (UV-killed) Bordetella avium and ferritin . Suspensions of bacteria and ferritin were instilled via intratracheal catheterization and allowed to remain in contact with the respiratory surfaces for 0, 10, 30, 60, 90, and 120 min . Ferritin and B . avium were taken up by both ciliated and non-ciliated cells of the epithelium overlying BALT (BALT epithelium) . Ferritin was found in organelles associated with endocytosis (i.e . apical vesicles, endosomes, cytoplasmic vacuoles) and was apparently transported across epithelial cells, since it was also found in intercellular spaces . Bacteria were found in vacuoles within BALT epithelial cells, but not free in intercellular spaces . Some macrophages in BALT epithelium also contained bacteria . No differences were observed between uptake of live and UV-killed bacteria . We conclude that both ciliated and non-ciliated cells of BALT epithelium in turkeys are able to take up macromolecular and particulate materials . Bacteria are also accessible to intraepithelial macrophages, although whether they are taken up directly from the bronchial surface or whether they pass through epithelial cells first could not be determined . This evidence suggests that antigens, including respiratory pathogens, could gain access to cells of the avian immune system by transepithelial passage in BALT. Vet Microbiol, 1994 Apr, 39(3-4), 205 - 18 NAD-independent Actinobacillus pleuropneumoniae strains: production of RTX toxins and interactions with porcine phagocytes; Dom P et al.; Actinobacillus pleuropneumoniae RTX toxin (Apx) production by A . pleuropneumoniae biotype 2 (NAD-independent) serotype 2 strains was studied . Western blot analysis of culture supernatants of all biotype 2 strains tested revealed the presence of a 103 kDa protein which reacted with a monoclonal antibody against ApxIIA . This protein was also recognized by sera of pigs infected with a biotype 2-serotype 2 strain . Furthermore, antibodies that could neutralize ApxIIA were present in these sera . Proteins corresponding to ApxIA or ApxIIIA were not detected . The effects of a biotype 1-serotype 2 and a biotype 2-serotype 2 strain and their metabolites on the oxidative activity of porcine pulmonary alveolar macrophages (PAM) and polymorphonuclear cells (PMN) were compared using a chemiluminescence (CL) technique . Viable bacteria of both biotypes stimulated the production of oxygen radicals by phagocytes . CL responses were higher for the biotype 1 than for the biotype 2 strain . After having reached a peak value, the oxidative activity decreased until a total inhibition was achieved . Inactivated washed bacteria had no influence on the oxidative activity of phagocytes . In contrast, heat labile factors in culture supernatants of both biotypes stimulated and inhibited the oxidative activity of PAM in a dose-dependent manner . Dilutions of supernatant up to 1/32 of the biotype 2 strain and up to 1/512 of the biotype 1 strain were toxic for PAM, while dilutions from 1/64 to 1/128 of the biotype 2 strain and from 1/1024 to 1/4096 of the biotype 1 strain stimulated the oxidative activity.(ABSTRACT TRUNCATED AT 250 WORDS) Eur J Clin Nutr, 1994 Apr, 48(4), 266 - 72 Reproducibility of the breath hydrogen measurement after a low and high fibre meal; Gelissen IC et al.; OBJECTIVE: To assess the intra-subject variability of the breath hydrogen (H2) response to a low and a high fibre test meal . DESIGN: Six week trial, consisting of three phases: screening (1 weeks), baseline period (1 week) and high fibre period (4 weeks) . SETTING: Western General Hospital, Edinburgh, Scotland . SUBJECTS: Sixteen subjects from the hospital staff and student population were screened for breath methane (CH4) production . Seven non-CH4 producers, four males and three females, were included in the trial . All completed the study successfully . INTERVENTION: Breath H2 and CH4 production were measured over a 12h period on duplicate test days after a low and high fibre meal . The high fibre meal was consumed for 4 weeks before the high fibre test days to allow for bacterial adaptation . The effect of the high fibre meal on stool output and whole gut transit time was furthermore assessed . RESULTS: Individual differences in breath H2 area-under-the-curve between duplicate test days ranged from 4% to 39% after the low fibre meal and from 4% to 37% after the high fibre meal . Considerable variation was found to be inherent in the breath collection method . In two subjects, breath CH4 was detected in response to the test meals . Stool output and whole gut transit time remained unchanged . CONCLUSION: Considerable differences were found between duplicate breath H2 responses to standard test meals . The variation inherent in the collection procedure emphasizes the need to collect breath samples at least in duplicate . The data presented here can be useful in future sample size calculations for similar studies. Curr Opin Genet Dev, 1994 Apr, 4(2), 221 - 8 Recombination genes and proteins; Dunderdale HJ et al.; The recombination of DNA takes place by a multistep process involving numerous gene products . In the past year, studies using bacterial proteins have led to a number of significant advances in our understanding of the enzymes of recombination and of the reactions that they catalyze . Moreover, the identification of eukaryotic proteins that are structurally analogous to the principal bacterial recombination enzyme, RecA protein, suggests that the basic mechanisms of homologous pairing and strand exchange have been conserved through evolution from bacteria to man. J Clin Microbiol, 1994 Apr, 32(4), 1043 - 9 Animal and public health implications of gastric colonization of cats by Helicobacter-like organisms; Otto G et al.; The bacterial genus Helicobacter contains a number of species which colonize the gastric mucosa of mammals . Natural and/or experimental gastric pathology has been correlated with colonization in humans and a wide variety of animal species . Historical reports in the literature suggest that a high percentage of cats are colonized by large, spiral, gastric helicobacter-like organisms (GHLOs) . One of these bacteria (Helicobacter felis) has been isolated on artificial media and has experimentally caused gastritis in gnotobiotic dogs . This study surveyed the prevalence of helicobacter colonization in random-source cats by using the urease assay . Histologic examination was performed to determine the degree of associated pathology present . GHLOs associated with chronic gastritis were present in 70% of the juvenile and 97% of the adult cats studied . Although further study is needed to determine specifically what role GHLOs play in feline gastrointestinal disease, these results indicate that helicobacter colonization should be considered in the pathogenesis of feline gastroenteropathy . Furthermore, the high prevalence of feline infection is interesting because cats have recently been implicated as a potential reservoir for human infection by helicobacter-like organisms. J Clin Microbiol, 1994 Apr, 32(4), 1018 - 22 A new method for identification of Trichomonas vaginalis by fluorescent DNA in situ hybridization; Muresu R et al.; The protozoan flagellate Trichomonas vaginalis is responsible for human trichomoniasis, one of the most widespread sexually transmitted diseases in the world . Several methods are currently used for laboratory diagnosis, including direct microscopic observation, cell culture, immunological techniques, and more recently, DNA probing and gene amplification . This report describes an in situ hybridization technique with specific DNA probes labeled with either biotin, rhodamine, or fluorescein for detection of T . vaginalis with fluorescence microscopy . Repetitive DNA sequences were evident in the nuclei of the protozoa as intensively fluorescent regions, giving a spotted pattern . No cross-reactivity between the probes and the DNAs of mammalian cells, yeasts, or bacteria was noted . This technique is potentially useful for the diagnosis of human trichomoniasis in clinical samples. Curr Opin Cell Biol, 1994 Apr, 6(2), 275 - 9 Salicylic acid as a signal molecule in plant-pathogen interactions; Vernooij B et al.; Significant insight has been gained in the past year into the roles of salicylic acid (SA) in plant-pathogen interactions . The ability to accumulate SA has been shown to be essential for systemic acquired resistance in tobacco plants . Further experiments have shown that SA is apparently not a systemic, vascular-mobile signal, but rather is required for signal transduction at the local level . Its mode of action may include inhibition of catalase activity, leading to increased levels of hydrogen peroxide. Artif Organs, 1994 Apr, 18(4), 318 - 21 Complications and side effects associated with large-bore catheters in the subclavian and internal jugular veins; Bambauer R et al.; Since the introduction of large-bore catheters for acute hemodialysis 30 years ago, many problems with handling, material, and contamination of these catheters exist . Nevertheless, catheterization of the inferior and superior vena cava with a large-bore catheter has proved to be suitable as a rapid connection process for hemodialysis, hemofiltration, hemoperfusion, plasmapheresis, plasmaperfusion, among others . In a retrospective study with 2,741 large-bore catheters in 1,716 patients, the frequency of infections, thrombosis, bleeding, and other side effects was investigated . All complications and side effects are presented dependent on vascular access route . In total, the complication rate was 48.9% higher in subclavian puncture than in internal jugular puncture (24.8%) . The highest complication rates for both vascular access routes were infections or septicemia; infections were observed in 19.5% of subclavian catheters versus 10% of internal jugular catheters. Artif Organs, 1994 Apr, 18(4), 272 - 5 Scanning electron microscopic investigation of catheters for blood access; Bambauer R et al.; Large-bore catheters for extracorporeal detoxification methods without and with treated surface with silver or silicone were investigated after removal with a scanning electron microscope and for bacterial colonization . In 42 large-bore catheters of three different materials, small deposits of fibrin and protein on the inner and outer surface were seen . This second layer covered the entire surface after 3 days and increased to a thickness of 3 to 60 microns during the following days . Bacterial colonization was observed in 38.1% . In contrast to these results, the catheters with the treated outer surface showed a very low thrombogenicity and a low contamination rate of 6.7%. Parasitology, 1994 Apr, 108 ( Pt 3), 289 - 300 Immunochemical studies on the cercarial-specific calcium binding protein of Schistosoma mansoni; Ram D et al.; Stage-specific expression of the mRNA encoding the cercarial-specific 8 kDa CaBP has been described previously . To gain information on possible function(s) of this protein we raised antibodies to the CaBP in rabbits immunized with a CaBP-TrpE fusion protein synthesized in bacteria . Western blots showed high levels of CaBP in cercariae and 3 h schistosomula, trace amounts in 24 h schistosomula, and none in miracidia sporocyst and adult worm, as found for the mRNA . The CaBP molecule has a short half-life (< or = 4 h) similar to that of the mRNA . Other experiments demonstrate that the CaBP may interact with a putative target molecule in a calcium-dependent manner to form a complex of 45 kDa . Immunogold electron microscopy showed CaBP in selected regions of cercariae and 3 h schistosomula: tegument, head gland, subtegumental cells, flame cells, intestinal wall and the body-tail junction . Other investigators have shown that the head gland and subtegumental cells synthesize and translocate granules to the tegument during transformation from cercariae (living freely in water) to schistosomula (residing in vertebrate host) . These observations and the time-course of CaBP detection suggest that the CaBP synthesized in the head gland and subtegumental cells is translocated to the tegument where it plays a role in tegument modifications required for adaptation to parasite life in the host . CaBP was not found in muscles and mitochondria, suggesting that it is not involved in the rapid motility and aerobic metabolism characteristic of cercariae. J Nat Prod, 1994 Apr, 57(4), 524 - 7 Structure of malhamensilipin A, an inhibitor of protein tyrosine kinase, from the cultured chrysophyte Poterioochromonas malhamensis; Chen JL et al.; A new chlorosulfolipid, malhamensilpin A {1} was isolated from the cultured chrysophyte Poterioochromonas malhamensis . Malhamensilipin A was demonstrated to be a modest inhibitor of pp60v-src protein tyrosine kinase . The structure was determined by detailed spectral analysis to be a novel C24 hexachloro lipid containing a vinyl sulfate ester (2,11,12,13,15,16-hexachloro-14-hydroxy-n-tetracos-1E-enol-1-sulfa te). Microsc Res Tech, 1994 Apr 1, 27(5), 376 - 88 Biomineralization: new directions in crystal science; Heywood BR; Effective protocols for controlling crystal structure, size, and morphology attract considerable interest given the requirement for particles of modal size and shape in many areas of materials fabrication and the importance of crystallochemical selectivity in determining the exploitable properties of inorganic solids . For this reason biomineralization merits particular attention since many biominerals are deposited in a highly controlled manner to produce crystals which are uniformly sized and crystallographically unique . Studies of biominerals have revealed that while a complex array of strategies have evolved for regulating their formation, one feature is common to the biological paradigm; interactions between organized biopolymeric assemblies and the nascent inorganic solids play a pivotal role in controlling the crystallization process . In order to gain a better understanding of the molecular interactions which take place at organic-inorganic interface and address the fundamental chemical problems of biomineralization, a crystal chemical approach has been adopted . Organized organic assemblies (phospholipid vesicles, Langmuir monolayers, polypetide templates) of precise molecular design (head group identity, packing conformation, peptide sequence, etc.) were assayed for their effectiveness in controlling the nucleation and growth of inorganic solids . This work has established that through systematic changes in the nature of the organic matrix the size, crystallographic orientation, and growth of the mineral phase can be controlled . Critical to this process was the translation of specific molecular information at the organic-inorganic interface: epitaxial alignment, stereochemical complementarity, and electrostatic interactions were an essential feature of this recognition event. Appl Environ Microbiol, 1994 Apr, 60(4), 1160 - 5 Seasonal and spatial population dynamics of the nitrogen-efficient guild in a desert bajada grassland; Herman RP et al.; This study examined the temporal and spatial variation in the populations of bacteria from a Chihuahuan desert bajada grassland that could grow on nearly nitrogen-free medium, a nitrogen-efficient guild (NEG) . A Curtis similarity index of 0.876 and nearly identical diversity and equability indexes (H' = 1.22, J = 0.46 and H' = 1.27, J = 0.48 for the crown edge and interplant samples, respectively) indicated that there was no qualitative difference between the NEG assemblages isolated from samples taken at Bouteloua eriopoda plant crowns and in nonvegetated areas 45 cm from crowns . The difference in NEG populations between the crown and interplant samples was only 2-fold, despite a 9.5-fold difference in root biomass between the sites . These differences, while consistent, were statistically significant at only 50% of the sampling times . There was over an order of magnitude difference in NEG numbers in root-associated soil and in bulk soil from the crown or intercrown sites . The typical trend for temporal variation in NEG numbers was that they increased in the spring, fluctuated dramatically over the summer, and declined at the summer's end . The pattern of soil moisture change was the only abiotic variable which showed the same fluctuation pattern as NEG numbers. Plant Physiol, 1994 Apr, 104(4), 1351 - 7 cDNA cloning of carrot (Daucus carota) soluble acid beta-fructofuranosidases and comparison with the cell wall isoenzyme; Unger C et al.; Carrot (Daucus carota), like most other plants, contains various isoenzymes of acid beta-fructofuranosidase (beta F) (invertase), which either accumulate as soluble polypeptides in the vacuole (isoenzymes I and II) or are ionically bound to the cell wall (extracellular beta F) . Using antibodies against isoenzyme I of carrot soluble beta F, we isolated several cDNA clones encoding polypeptides with sequences characteristic of beta Fs, from bacteria, yeast, and plants . The cDNA-derived polypeptide of one of the clones contains all partial peptide sequences of the purified isoenzyme I and thus codes for soluble acid beta F isoenzyme I . A second clone codes for a related polypeptide (63% identity and 77% similarity) with characteristics of isoenzyme II . These two soluble beta Fs, have acidic isoelectric points (3.8 and 5.7, respectively) clearly different from the extracellular enzyme, which has a basic isoelectric point of 9.9 . Marked differences among the three nucleotide sequences as well as different hybridization patterns on genomic DNA gel blots prove that these three isoenzymes of carrot acid beta F are encoded by different genes and do not originate from differential splicing of a common gene, as is the case in the yeast Saccharomyces cerevisiae . All three carrot acid beta Fs, are preproenzymes with signal peptides and N-terminal propeptides . A comparison of the sequences of the soluble enzymes with the sequence of the extracellular protein identified C-terminal extensions with short hydrophobic amino acid stretches that may contain the information for vacuolar targeting. APMIS, 1994 Apr, 102(4), 291 - 4 Bordetella pertussis diagnosed by polymerase chain reaction; Birkebaek NH et al.; The object of this work was to test the polymerase chain reaction (PCR) for demonstration of Bordetella pertussis (BP) in nasopharyngeal secretions . The method was applied to patients with recently diagnosed pertussis, as verified by BP culture . In order to test the sensitivity and specificity of PCR for the diagnosis of BP, we used known concentrations of BP, Bordetella parapertussis and Bordetella bronchiseptica in aqueous solutions . PCR was furthermore carried out on species of bacteria that might be isolated from the nasopharynx . The applicability of PCR to patient specimens was tested in 25 patients in whose nasopharyngeal secretions BP had been demonstrated after 4-7 days of culture . The detection limit of PCR in aqueous solution was 1-2 BP bacteria per reaction tube . PCR was 100% specific for BP, showing no response with other Bordetella species or other bacteria known to colonize the nasopharynx . Of 25 patient specimens, 16 were PCR-positive 4-7 days after the positive primary culture had been established; only 5 out of 13 patient specimens were positive by repeated conventional nasopharyngeal culture at that time . We conclude that PCR is a possible alternative to culture for the demonstration of BP, as PCR is considerably faster than culture and might be more sensitive. Zentralbl Hyg Umweltmed, 1994 Apr, 195(4), 306 - 18 {Ventilation method plan in daily operations--a practical study}; Bischoff WE et al.; On three days each we measured particles and airborne bacteria in ten ventilated operating theatres . Modern air conditioning systems achieved a significant reduction of air-pollution . Vertical systems proved to be more effective than horizontal systems . Workday conditions made several problems: During high personal activity a body exhaust system was not able to reduce the concentration of airborne bacteria; doors near the operating table and objects in the airstream had a negative effect; there was more personal than necessary in the operating theatres; some activities took place without a specific purpose and resulted in raised particle and bacteria contamination; early clearing of materials, before the operation ended, increased turbulences and door movements . A short break in personal activity before the first skin cut is recommended to reduce high air contamination due to the preparation of the operating room and the patient . One of the ventilation systems was insufficiently operated by personal . We recommend continuous measurement of particles near the operating field in order to control the input of airborne particles and bacteria into the wound. J Mol Evol, 1994 Apr, 38(4), 319 - 27 The evolutionary origin of red algae as deduced from the nuclear genes encoding cytosolic and chloroplast glyceraldehyde-3-phosphate dehydrogenases from Chondrus crispus; Liaud MF et al.; Algae are a heterogeneous group of photosynthetic eukaryotes traditionally separated into three major subdivisions: rhodophytes, chlorophytes, and chromophytes . The evolutionary origin of rhodophytes or red algae and their links to other photosynthetic and nonphotosynthetic eukaryotes have been a matter of much controversy and speculation . Here we present the first cDNAs of nuclear protein genes from red algae: Those encoding cytosolic and chloroplast glyceraldehyde-3-phosphate dehydrogenases (GAPDH) from Chondrus crispus . A phylogenetic analysis including GAPDH gene sequences from a number of eukaryotic taxa, cyanobacteria, and purple bacteria suggests that chloroplasts and rhodoplasts together form a monophyletic group of cyanobacterial descent and that rhodophytes separated from chlorophytes at about the same time as animals and fungi . The composite GAPDH tree further demonstrates that chloroplast and cytosolic GAPDH genes are closely related to their homologs in cyanobacteria and purple bacteria, respectively, the presumptive ancestors of chloroplasts and mitochondria, thereby firmly establishing the endosymbiotic origin of these nuclear genes and their fixation in eukaryotic cells before the rhodophyte/chlorophyte separation . The present data are in conflict with phylogenetic inferences based on plastid-encoded rbcL sequences supporting a polyphyletic origin of rhodoplasts and chloroplasts . Comparison of rbcL to GAPDH phylogenies suggests that rbcL trees may be misleading because they are composed of branches representing ancient duplicated (paralogous) genes. Avian Dis, 1994 Apr-Jun, 38(2), 376 - 8 Clinical and pathological findings in young Georgia broiler chickens with oculofacial respiratory disease ("so-called swollen heads"); Goodwin MA et al.; Chickens with swollen heads generally are said to have swollen head syndrome . In this retrospective study, 16 accessions of young chickens during 1992 had "swollen heads." Clinical signs and lesions were accompanied by infection with multiple viruses, bacteria, and Cryptosporidium baileyi . The fact that almost half of the cases of chickens with swollen heads occurred in one broiler-producing company and predictably within 14 days post-vaccination suggests that management factors might be instrumental in inducing the incidence of swollen heads. Microb Pathog, 1994 Apr, 16(4), 261 - 7 Borrelia burgdorferi decreases hyaluronan synthesis but increases IL-6 production by fibroblasts; Jones NC et al.; Despite the prevalence of clinical data on human Lyme disease, little is known about the immunopathologic effects of the causative organism on the host . We studied the effect of Borrelia burgdorferi on hyaluronan (hyaluronic acid, HYA) production and the effect on interleukin-6 (IL-6) synthesis by cultured fibroblasts . The cell line employed in this study produced an average of 1406 ng of hyaluronan/ml within 48 h . Using both a morphological staining protocol and a quantitative radiometric assay, we noted that in the presence of a low dose of Borrelia (9.4 x 10(5) cells/ml) the hyaluronan production decreased to an average of 1008 ng/ml, a significant difference (p < 0.05) from the amount of hyaluronan produced by the cells alone . The reduction was even more significant (p < 0.01) when a higher dose of Borrelia (9.4 x 10(6) cells/ml) was used giving an average hyaluronan concentration of 682 ng/ml . In contrast, we found that Borrelia stimulated the cells to produce IL-6 from a baseline of 293 pg/ml to a maximal value of 842 pg/ml (p < 0.01) . The spirochetes had no significant effect on cell viability, nor were we able to demonstrate invasion of the cells by the bacteria . Both a decrease in hyaluronan and an increase in IL-6 may correlate with the pathogenicity of Lyme disease in man. Infect Immun, 1994 Apr, 62(4), 1262 - 7 In vivo association of Actinobacillus pleuropneumoniae serotype 2 with the respiratory epithelium of pigs; Dom P et al.; The ability of an Actinobacillus pleuropneumoniae serotype 2 strain to associate in vivo with the epithelium of the porcine respiratory tract was investigated in a sequential study after intranasal inoculation of hysterectomy-derived and colostrum-deprived pigs . At 30 min postinoculation more than 95% of the bacteria present in the lungs were intimately associated with the epithelium of the alveoli or the cilia of the terminal bronchioli, as observed by light and electron microscopy . At 90 and 180 min postinoculation multiple focal early inflammatory lesions in which histologically different, more or less concentric zones could be distinguished were observed . In the center of these pneumonic areas bacteria were associated with infiltrated cells and exudate . In the zone surrounding the center, approximately 95% of the bacteria were lying with their longest side in close apposition to the epithelial cells of alveoli and the cilia of the terminal bronchioli . Bacteria were only sporadically associated with the cilia or the epithelium of the bronchi and trachea . Bacteria were not observed in tonsils or conchae . In view of the findings presented here, we propose the hypothesis that adherence of the A . pleuropneumoniae serotype 2 strain to epithelial cells of the lower respiratory tract constitutes an important initial step in pathogenesis. Dent Update, 1994 Apr, 21(3), 103 - 6 Erosion in children: an increasing clinical problem? Shaw L, Smith A. Tooth wear is becoming more common in both adults and children . The triad of attrition, abrasion and erosion has been known for many years, but the contribution of erosion (irreversible loss of dental hard tissue due to a chemical process not involving bacteria, or the loss of tooth surface not directly associated with mechanical or traumatic factors or caries) to excessive loss of tooth tissue is now being emphasized . The authors of this paper examine the problem and suggest ways of overcoming it. Indian J Public Health, 1994 Apr-Jun, 38(2), 44 - 9 Usefulness of ORT in certain special situations of diarrhoeal diseases; Dutta P; PIP: Diarrhea is one of the most common causes of morbidity and mortality in infants and children less than 5 years old in developing countries . Diarrheal diseases are a major cause of childhood malnutrition . Toxin-producing bacteria are responsible for many acute diarrheas . Oral rehydration solution (ORS) treats dehydration caused by acute diarrheal episodes . WHO promotes the use of a single oral rehydration formula which contains 3.5 g sodium chloride, 2.5 g sodium bicarbonate or 2.9 g trisodium citrate dihydrate, 1.5 g potassium chloride, and 20 g glucose to 1 liter of water . This ORS formula can safely be used for all age groups and all etiologies of diarrhea . ORS replaces the lost fluid and electrolytes and maintains fluid and electrolytes . Pediatricians in most developed countries do not accept this ORS formula in cases of rotavirus-caused diarrhea because rotavirus blunts some absorptive villi and reduces the activity of lactase and other disaccharidase, resulting in reduced absorption . Yet, the unaffected villus cells may absorb enough water and electrolytes to be effective . In cases of vomiting, ORS should be administered in small amounts and slowly . Some health workers are concerned that 90 mmol/l sodium in the WHO formula causes hypernatremia in neonates and young infants who have low sodium levels in their stools . Specialists suggest ORS with 30-60 mmol/l or additional water administered in a 2:1 ratio for these young infants . Hypernatremia is also a concern for malnourished children, but studies show that WHO's ORS is safe and effective in treating malnourished children . Bottle fed children are more vulnerable to hypernatremia than breast fed children . Hypernatremia has neurological effects . Hyponatremia is more common in developing countries than developed countries . It also has neurological effects . In severe dehydration cases, intravenous fluid or ORS delivered via a nasogastric tube should be given immediately . Bioseparation, 1994 Apr, 4(2), 73 - 83 Ultrasonic manipulation of particles and cells . Ultrasonic separation of cells; Coakley WT et al.; Cells or particles suspended in a sonic standing wave field experience forces which concentrate them at positions separated by half a wavelength . The aims of the study were: (1) To optimise conditions and test theoretical predictions for ultrasonic concentration and separation of particles or cells . (2) To investigate the scale-up of experimental systems . (3) To establish the maximum acoustic pressure to which a suspension might be exposed without inducing order-disrupting cavitation . (4) To compare the efficiencies of techniques for harvesting concentrated particles . The primary outcomes were: (1) To design of an acoustic pressure distribution within cylindrical containers which led to uniformly repeating sound pressure patterns throughout the containers in the standing wave mode, concentrated suspended eukaryotic cells or latex beads in clumps on the axis of wide containers, and provided uniform response of all particle clumps to acoustic harvesting regimes . Theory for the behaviour (e.g . movement to different preferred sites) of particles as a function of specific gravity and compressibility in containers of different lateral dimensions was extended and was confirmed experimentally . Convective streaming in the container was identified as a variable requiring control in the manipulation of particles of 1 micron or smaller size . (2) Consideration of scale-up from the model 10 ml volume led to the conclusion that flow systems in intermediate volume containers have more promise than scaled up batch systems . (3) The maximum acoustic pressures applicable to a suspension without inducing order-disrupting cavitation or excessive conductive streaming at 1 MHz and 3 MHz induce a force equivalent to a centrifugal field of about 10(3) g . (4) The most efficient technique for harvesting concentrated particles was the introduction of a frequency increment between two transducers to form a slowly sweeping pseudo-standing wave . The attractive inter-droplet ultrasonic standing wave force was employed to enhance the rate of aqueous biphasic cell separation and harvesting . The results help clarify the particle size, concentration, density and compressibility for which standing wave separation techniques can contribute either on a process engineering scale or on the scale of the manipulation of small particles for industrial and medical diagnostic procedures. J Cell Biol, 1994 Apr, 125(2), 483 - 93 The distribution of tenascin-X is distinct and often reciprocal to that of tenascin-C; Matsumoto K et al.; We have isolated a cDNA encoding mouse tenascin-X (TN-X), a new member of the family of tenascin genes . The TN-X gene lies in the major histocompatibility complex (MHC) class III region, as it is the case for its human counterpart . On Northern blots we detected a TN-X mRNA of approximately 13 kb in most tissues analyzed, whereas in various mouse cell lines mRNAs of approximately 11 and 13 kb were detected, suggesting the possibility of alternative splicing of TN-X transcripts . We raised antibodies against mouse TN-X fragments expressed in bacteria and used these antibodies to identify the TN-X protein in heart cell extracts and in the conditioned medium of a renal carcinoma cell line . The subunit molecular size of TN-X is approximately 500 kD, suggesting that the protein may contain up to 40 fibronectin type III repeats, making it the largest tenascin family member known yet . TN-X in conditioned medium, as well as the purified protein bind to heparin, but no binding to tenascin-C (TN-C), fibronectin, laminin or collagens could be detected . Thus the heparin-binding activity may be a common feature of the tenascins . The TN-X mRNA as well as the protein are predominantly expressed in heart and skeletal muscle, but the mRNA is found in most tissues at a low level . Immunostaining showed the protein to be associated with the extracellular matrix of the muscle tissues and with blood vessels in all of the tissues analyzed . Although the TN-X gene lies in the MHC class III locus, it is not expressed in the lymphoid organs analyzed, except for the staining around blood vessels . In skin and tissues of the digestive tract often a reciprocal distribution of TN-X and TN-C was observed. J Clin Endocrinol Metab, 1994 Apr, 78(4), 944 - 9 Recombinant thyroid peroxidase autoantibodies can be used for epitopic "fingerprinting" of thyroid peroxidase autoantibodies in the sera of individual patients; Nishikawa T et al.; Four human monoclonal antibodies (SP1.4, WR1.7, TR1.8, and TR1.9) map the immunodominant region on thyroid peroxidase (TPO) recognized by autoantibodies in patients' sera . We used a pool of these monoclonal antibodies, expressed in bacteria as antigen-binding fragments {F(ab)}, to compete for TPO autoantibody binding to radiolabeled TPO . The F(ab) inhibited TPO binding by 32 patients' sera by 82 +/- 14% (mean +/- SD), with a range from 51-100% . When each F(ab) was tested individually for its ability to compete for autoantibody binding to TPO, F(ab) TR1.8 was the most potent among the 32 sera . However, there was a wide spectrum of TPO binding inhibition when each serum was considered individually, thereby allowing an epitopic "fingerprint" to be drawn for the TPO autoantibodies in a patient's serum . There was a close association between the proportions of TPO autoantibodies to the TR1.8 and TR1.9 epitopes as well as between those to the SP1.4 and WR1.7 epitopes . These associations correspond to the previously described A and B epitopic domains in the TPO immunodominant region . No TPO epitope was observed to be associated with clinically apparent ophthalmopathy of Graves' disease, nor was there an association between TPO epitopes and patient age or sex . In summary, the present study on a large sample of sera with TPO autoantibodies indicates that by using TPO-specific F(ab) selected to cover all regions of the TPO immunodominant region, it is possible to obtain a TPO epitopic fingerprint for each serum . These data open the way to future studies directed at testing the hypothesis of disease-associated TPO epitope(s). Mutat Res, 1994 Apr, 317(2), 145 - 62 Potential genotoxic, mutagenic and antimutagenic effects of coffee: a review; Nehlig A et al.; Coffee and caffeine are mutagenic to bacteria and fungi, and in high concentrations they are also mutagenic to mammalian cells in culture . However, the mutagenic effects of coffee disappear when bacteria or mammalian cells are cultured in the presence of liver extracts which contain detoxifying enzymes . In vivo, coffee and caffeine are devoid of mutagenic effects . Coffee and caffeine are able to interact with many other mutagens and their effects are synergistic with X-rays, ultraviolet light and some chemical agents . Caffeine seems to potentiate rather than to induce chromosomal aberrations and also to transform sublethal damage of mutagenic agents into lethal damage . Conversely, coffee and caffeine are also able to inhibit the mutagenic effects of numerous chemicals . These antimutagenic effects depend on the time of administration of coffee as compared to the acting time of the mutagenic agent . In that case, caffeine seems to be able to restore the normal cycle of mitosis and phosphorylation in irradiated cells . Finally, the potential genotoxic and mutagenic effects of the most important constituents of coffee are reviewed . Mutagenicity of caffeine is mainly attributed to chemically reactive components such as aliphatic dicarbonyls . The latter compounds, formed during the roasting process, are mutagenic to bacteria but less to mammalian cells . Hydrogen peroxide is not very active but seems to considerably enhance mutagenic properties of methylglyoxal . Phenolic compounds are not mutagenic but rather anticarcinogenic . Benzopyrene and mutagens formed during pyrolysis are not mutagenic whereas roasting of coffee beans at high temperature generates mutagenic heterocyclic amines . In conclusion, the mutagenic potential of coffee and caffeine has been demonstrated in lower organisms, but usually at doses several orders of magnitude greater than the estimated lethal dose for caffeine in humans . Therefore, the chances of coffee and caffeine consumption in moderate to normal amounts to induce mutagenic effects in humans are almost nonexistent. Mol Cell Biochem, 1994 Mar 30, 132(2), 101 - 8 Immunological approach to identify calmodulin-stimulated phosphatase isozymes from bovine brain; Yokoyama N et al.; Molecular cloning of human, mouse and rat brain CaM-stimulated phosphatase has suggested the existence of two genes for the alpha subunit of the enzymes . A alpha and A beta fragments of A alpha and A beta from rat brain library have been expressed in bacteria to produce specific anti-calcineurin A alpha and anti-calcineurin A beta antibodies (Kuno et al., J Neurochem 58: 1643-1651, 1992) . Alternative mRNA splicing gives rise to additional calcineurin isozymes with some containing an insertion sequence of ATVEAIEADE . Antibody against synthetic peptide of this insertion sequence has been raised in this study . Three CaM-stimulated phosphatase isozymes previously purified from bovine brain (BPI, BPII, BPIII) (Yokoyama & Wang, J Biol Chem 266: 14822-14829, 1991), along with the bacterially expressed rat A alpha and A beta fragments, were analyzed by two calcineurin alpha subunit monoclonal antibodies VJ6 and VD3, the rat anti-calcineurin A alpha and anti-calcineurin A beta specific polyclonal antibodies, and the insertion peptide antibody . The bovine brain CaM-stimulated phosphatase isozymes BPI and BPIII reacted with both anti-calcineurin A alpha and anti-calcineurin A beta antibodies . While BPII reacted with anti-calcineurin A alpha but not anti-calcineurin A beta antibody, it differed from the expressed A alpha fragment in immunoreactivity towards the monoclonal antibodies . The results show that the bovine brain CaM-stimulated phosphatase isozymes cannot be simply categorized as derived from A alpha or A beta genes products.(ABSTRACT TRUNCATED AT 250 WORDS) Biochemistry, 1994 Mar 29, 33(12), 3572 - 6 Resonance Raman spectroscopic evidence for the FeS4 and Fe-O-Fe sites in rubrerythrin from Desulfovibrio vulgaris; Dave BC et al.; Resonance Raman (RR) spectra of the non-heme iron protein rubrerythrin from Desulfovibrio vulgaris unequivocally demonstrate the presence of both a rubredoxin-type FeS4 site and a (mu-oxo)diiron(III) cluster . The RR spectra of rubrerythrin excited at 496.5 and 568.2 nm are dominated by bands similar to those of rubredoxin and conform to the vibrational pattern expected for a distorted FeS4 tetrahedron of an Fe(S-Cys)4 site . Numerous overtone and combination bands of the Fe-S stretches are also observed, and a band at 650 cm-1 is assigned to a cysteine C-S stretching mode . The 374-, 355-, and 340-cm-1 bands, assigned to the three components of the v3(T2) asymmetric FeS4 stretching mode, are 2-8 cm-1 lower than the corresponding frequencies for the Desulfovibrio gigas rubredoxin FeS4 site . Similar differences in frequencies of bands assigned to SFeS bending modes between rubredoxin and rubrerythrin are also detected . These frequency differences imply either slightly weaker Fe-S bonds or subtle conformational differences among the cysteinyl ligands in the rubrerythrin versus rubredoxin FeS4 sites . The RR spectrum of rubrerythrin excited at 406.7 nm shows dramatically diminished intensities of the FeS4 bands with concomitant enhancement of a band at 514 cm-1 . This band shifts 18 cm-1 to lower frequency when the protein is dissolved in H(2)18O . The frequency of this band and the 18O isotope shift are those expected for the symmetric Fe-O-Fe stretch of a bent oxo-bridged diiron(III) cluster and indicate that this cluster has at least one additional bridging ligand.(ABSTRACT TRUNCATED AT 250 WORDS) FEBS Lett, 1994 Mar 28, 342(1), 61 - 5 The primary structure of two chlorosome proteins from Chloroflexus aurantiacus; Niedermeier G et al.; The complete nucleotide sequence of two chlorosome proteins with apparent molecular weights of M(r) 18,000 and M(r) 11,000 from Chloroflexus aurantiacus have been determined . The two polypeptides were 145 and 97 amino acids long and possessed true molecular masses of 15,545 and 10,820 Da, respectively . Protein chemical sequencing was done in parallel to confirm the primary structure deduced from nucleotide sequencing . By Northern blot analysis of RNA isolated from phototrophically grown cells a transcript of 0.95 kb was detected which is the expected length for a mRNA encoding both genes. Gene, 1994 Mar 25, 140(2), 273 - 8 Genomic cloning of human thioredoxin-encoding gene: mapping of the transcription start point and analysis of the promoter; Kaghad M et al.; Thioredoxin (TR) is a small ubiquitous dithiol-reductase enzyme first identified in bacteria and plants . In recent years, this protein has been recognized as playing an important role in the growth control of eukaryotic cells, especially in lymphocytes . It was first cloned from a human Epstein-Barr virus-transformed lymphoblastoid B-cell line by our group in 1988 {Wollman et al., J . Biol . Chem . 263 (1988) 15506-15512} and localized on chromosome 3 p11-p12 by in situ hybridization {Lafage-Pochitaloff-Huvale et al., FEBS Lett . 255 (1989) 89-91} . The present work was performed to study the genomic organization of the human thioredoxin (hTR)-encoding gene (hTR) . The screening of a human genomic library in lambda EMBL4 phage led to the identification of two genomic clones which encompassed the entire gene, including the promoter region . The coding region of hTR spans over 13 kb and is organized into five exons separated by four introns which were 60% sequenced . We determined the transcription start point (tsp) by primer extension . This tsp located, in lymphocytes, 22-bp downstream from a TATA box (TATAA) defines a 5' untranslated region of 74 bp . We analyzed 2149-bp upstream from the promoter for sequence motifs which could bind regulatory proteins . This promoter contains many possible regulatory elements compatible with both a basal constitutive expression and a regulated inducible transcription, especially by cytokines such as interleukin-6 and interferons . Finally, Southern hybridization of genomic DNAs from several donors detected only one active gene encoding hTR. J Biol Chem, 1994 Mar 25, 269(12), 9374 - 9 Mutations in the zinc-finger region of the yeast regulatory protein ADR1 affect both DNA binding and transcriptional activation; Cook WJ et al.; The expression of the yeast ADH2 gene is controlled by the transcriptional activator ADR1, a zinc-finger protein that binds to an upstream activating sequence (UAS1) in the ADH2 promoter . We report here the isolation of seven mutations in the ADR1-5c allele, defining five different amino acid changes, that suppress the enhanced ADH2 expression caused by the ADR1-5c allele . Each of the mutations was shown to reduce the activation of ADH2 by a wild-type ADR1 gene, suggesting the mutations disrupt a domain important to the function of both the ADR1 and ADR1-5c proteins . All five amino acid changes occurred within the DNA-binding domain of ADR1 and were shown to severely inhibit the ability of ADR1 to bind UAS1 in vitro . These mutations were found, however, to also affect the ability of ADR1 to activate transcription independent of its ability to bind DNA . These results indicate that the DNA-binding region of ADR1 is involved in both transactivation and DNA binding. J Biol Chem, 1994 Mar 25, 269(12), 8653 - 8 The carboxyl-terminal domain of lipoprotein lipase binds to the low density lipoprotein receptor-related protein/alpha 2-macroglobulin receptor (LRP) and mediates binding of normal very low density lipoproteins to LRP; Williams SE et al.; Lipoprotein lipase (LPL) binds with high affinity to the low density lipoprotein receptor-related protein/alpha 2-macroglobulin receptor (LRP) and promotes binding, uptake, and degradation of normal triglyceride-rich lipoproteins in a process mediated by LRP (Chappell, D . A., Fry, G . L., Naknitx, M.A., Muhonen, L . E., Pladet, M . W., Iverius, P-H., and Strickland, D . K . (1993) J . Biol . Chem . 268, 14168-14175) . To localize the portion of LPL that is responsible for interacting with LRP, fragments of LPL were expressed in bacteria . A fragment of human LPL containing the COOH-terminal domain (residues 313-448, designated LPLC) which lacks the catalytic site was able to bind to LRP . Purified LRP bound specifically to microtiter wells coated with LPL or LPLC with KD values of 2.8 and 5 nM, respectively . The effects of several mutations of LPLC were tested . Mutation of Lys407 to Ala reduced the affinity of LPLC for LRP by approximately 10-fold . Like native LPL, LPLC prevented the binding of activated alpha 2-macroglobulin and the 39-kDa receptor-associated protein to LRP and inhibited the internalization and degradation of activated alpha 2-macroglobulin and receptor-associated protein in cultured fibroblasts . LPLC also bound to 125I-labeled human normal triglyceride-rich lipoproteins and promoted their binding to purified LRP and to cultured cells . Mutation of Trp393 and Trp394 to Ala completely abolished the ability of LPLC to bind to lipoproteins, but had little effect on its interaction with LRP . These data indicate that the COOH-terminal domain of LPL may function both in binding lipoproteins and mediating their interaction with LRP. Biochim Biophys Acta, 1994 Mar 24, 1211(3), 243 - 55 Structure and biosynthesis of marine algal oxylipins; Gerwick WH; Diverse marine life, including algae, sponges, molluscs, corals, tunicates, and bacteria, have been found to possess a variety of structurally unique oxylipins . The algae are the best characterized of these organisms for their oxylipins, which have now been described from more than 30 species representing the three major groups of macrophytic algae (Rhodophyta = reds, Chlorophyta = greens, and Phaeophyceae = browns) . A number of recent studies have sought to understand the biosynthetic origin and mechanistic chemistry which leads to the formation of these unique marine substances . In general, the red algae metabolize C20 acids via 12-lipoxygenase-initiated pathways, green algae metabolize C18 acids at C-9 and C-13, and brown algae metabolize both C18 and C20 acids, principally by lipoxygenases with n-6 specificity . This review updates the records of new oxylipins from marine algae and describes thoughts on their biogenesis as well as specific experiments aimed at probing these hypotheses. Med J Aust, 1994 Mar 21, 160(6), 358 - 63,366 Sexually transmitted diseases in the tropics; Perine PL; Several sexually transmitted diseases are endemic in the tropics . The morbidity and mortality from the human immunodeficiency viruses (HIV-1 and HIV-2) alone now rival that caused by Plasmodium falciparum malaria in several African and Asian nations . The genital ulcers of chancroid and syphilis facilitate the sexual transmission of HIV . Within the last two decades, the bacteria causing chancroid and gonorrhoea throughout the world have acquired plasmids that mediate bacterial resistance to penicillins and other antibiotics . This has significantly increased the costs of treatment . There is little prospect that the prevalence of gonorrhoea, chancroid, syphilis and HIV will decrease in the tropics in the near future without a global change in sexual behaviours and practices. J Biol Chem, 1994 Mar 18, 269(11), 8564 - 75 Azotobacter vinelandii ferredoxin I . Alteration of individual surface charges and the {4FE-4S}2+/+ cluster reduction potential; Shen B et al.; The structures of Azotobacter vinelandii ferredoxin I (AvFdI) and Peptococcus aerogenes ferredoxin (PaFd), near their analogous {4e-4S}2+/+ clusters, are highly conserved (Backes, G., Mino, Y., Loehr, T.M., Meyer, T.E., Cusanovich, M.A., Sweeney, W.V., Adman, E.T., and Sanders-Loehr, J . (1991) J . Am . Chem . Soc . 11, 2055-2064) . Despite these similarities, the reduction potential (E0') of the AvFdI {4Fe-4S}2+/+ cluster is more than 200 mV more negative than that of PaFd . We have tested the contribution that individual amino acid residues make to the control of E0' by converting residues in AvFdI into the corresponding residue in PaFd . Four mutations involved substitutions of negatively charged surface residues with neutral residues and two involved substitution of buried hydrophobic residues . All AvFdI variants were characterized by x-ray crystallography, absorption, CD, EPR, and 1H NMR spectroscopies and by electrochemical methods . For the F25I mutation, significant structural changes occurred that affected the EPR and 1H NMR spectroscopic properties of AvFdI and had a minor influence on E0' . For all other mutations there were no changes in reduction potential . Thus we conclude, that variations in charged surface residues do not account for the observed differences in E0' between the analogous {4Fe-4S}2+/+ cluster of PaFd and AvFdI . These differences are therefore most likely to be due to differences in solvent accessibility. FEMS Microbiol Lett, 1994 Mar 15, 117(1), 107 - 11 Degradation of poly(3-hydroxybutyrate-co-3-hydroxyvalerate) by aerobic sewage sludge; Briese BH et al.; The degradation of sheets of poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (BIOPOL) by aerobic sewage sludge was analyzed . Degradation of the polymer was highly dependent on the pH of the culture medium and was maximal between pH 7 and pH 8.5 . Below pH 6 and above pH 9 the degradation rate was very low . Agitation of the culture fluid had relatively little influence on the rates of degradation . 1.2 x 10(5) aerobic polymer-degrading bacteria per ml sewage sludge were identified by halo formation on solid poly(3-hydroxybutyrate) (PHB)-containing media . The number of PHB-degrading bacteria in other ecosystems amounted to 3.8 x 10(3) per ml sludge of a fresh-water lake, 9.2 x 10(5) per g garden-soil, 1.3 x 10(6) per g field-soil and 4.3 x 10(6) per g compost. J Immunol, 1994 Mar 15, 152(6), 2720 - 8 Antigen presentation in the central nervous system . The inhibitory effect of IL-10 on MHC class II expression and production of cytokines depends on the inducing signals and the type of cell analyzed; Frei K et al.; Cells of the macrophage lineage are required to cope with bacterial infection and to serve as APC for T lymphocytes . Among the regulatory factors limiting the macrophage response to infection and the expansion of Ag-specific T cells, IL-10 has received recent attention . On monocytes/macrophages, IL-10 has been shown to inhibit the intracellular killing of bacteria, the secretion of cytokines, and the expression of MHC molecules . In the present study we have examined the effect of IL-10 on different APC obtained from the central nervous system . Both, astrocytes and microglial cells are in a resting state and require activation signals to express MHC class II and cytokine genes . Whereas IL-10 profoundly inhibits the IFN-gamma-induced expression of MHC class II Ag on microglial cells, it had no such effects on astrocytes . Nevertheless, IL-10 suppressed the MHC class II- and Ag-dependent proliferative response of T cells in the presence of both types of APC . As shown by the use of anti-IL-10 Abs, endogenously produced IL-10 influenced the function of microglia but not of astrocytes to serve as APC . IL-10 significantly inhibited the LPS-induced production of granulocyte-macrophage-CSF, macrophage-CSF, and IL-6 by both astrocytes and microglial cells . In contrast, the secretion of these cytokines by the two glial cell population was not altered by IL-10 when IL-1 beta, TNF-alpha, or viruses were used as stimuli. Eur J Biochem, 1994 Mar 15, 220(3), 901 - 10 Molecular cloning and sequence analysis of the gene of the molybdenum-containing aldehyde oxido-reductase of Desulfovibrio gigas . The deduced amino acid sequence shows similarity to xanthine dehydrogenase; Thoenes U et al.; In this report, we describe the isolation of a 4020-bp genomic PstI fragment of Desulfovibrio gigas harboring the aldehyde oxido-reductase gene . The aldehyde oxido-reductase gene spans 2718 bp of genomic DNA and codes for a protein with 906 residues . The protein sequence shows an average 52% (+/- 1.5%) similarity to xanthine dehydrogenase from different organisms . The codon usage of the aldehyde oxidoreductase is almost identical to a calculated codon usage of the Desulfovibrio bacteria. Biochem Biophys Res Commun, 1994 Mar 15, 199(2), 1027 - 34 Induction of thyrotropin receptor (TSH-R) autoantibodies and thyroiditis in mice immunised with the recombinant TSH-R; Costagliola S et al.; Two groups of 10 Balbc by Jico male mice were immunised on days 0, 15 and 35, with the extra cellular domain (ECD) of the human thyrotropin receptor (TSH-R) expressed as a fusion protein in bacteria (group 1) or with the maltose binding protein (MBP) fusion partner alone (group 2) . Blood was obtained on days 0, 22, 32, 42 and 49 and samples from the individual animals pooled for each group . Serum and immunoglobulin (IgG) preparations were tested, using CHO cells expressing the human TSH-R (JP26 and JP09) for thyroid stimulating (TSAb); thyroid blocking (TBAb) and thyrotropin binding inhibiting (TBII) activities . Neither serum nor IgGs were found to contain TSAb at any time point . TBII activity was present in the serum of both groups on day 32 and in group 1 only on day 49; when the test was performed on IgGs, only the MBP-ECD day 49 preparation remained significantly positive for TBII (p < 0.005) . Significant TBAb activity was present in both the serum and IgG of group 1 day 49 (p < 0.005) and to a lesser extent on 42 (p < 0.02) . Following the second immunisation (day 15) both groups and had decreased circulating T4 levels (p < 0.05) when compared with day 0 in each case . Group 2 were unaffected by the third immunisation on day 35 but the MBP-ECD group again had significantly decreased T4 levels (p < 0.02) compared with MBP day 49 and (p < 0.03) when compared with MBP-ECD day 0 . Histological examination of thyroids from group 1 animals revealed extensive vascularisation and an atypical lymphoblastoid infiltration which was not observed in control mice . These preliminary results indicate that care is required in interpreting data since a non-receptor antigen was shown to decrease circulating thyroxine and serum from these animals had apparent TBII like activity . However, the results obtained with the IgGs suggest that receptor autoantibodies can be induced by immunising with the human TSH-R, in addition, the immunised mice show histological evidence for the development of thyroiditis. Biochemistry, 1994 Mar 15, 33(10), 3113 - 9 A novel cytochrome c oxidase from Rhodobacter sphaeroides that lacks CuA; Garcia-Horsman JA et al.; Rhodobacter sphaeroides contains at least two different cytochrome c oxidases . When these bacteria are grown with high aeration, the traditional aa3-type cytochrome c oxidase is present at relatively high levels . However, under microaerophilic growth conditions or when the bacteria are grown photosynthetically, the amount of the aa3-type oxidase is greatly diminished and an alternate cytochrome c oxidase is evident . This alternate oxidase has been purified and characterized . The enzyme consists of three subunits by SDS-PAGE analysis (Mapp 45, 35, and 29 kDa) . Two of the three subunits (Mapp 35 and 29 kDa) contain covalently bound heme C . Metal and heme analyses indicate that the oxidase contains heme C, heme B (protoheme IX), and Cu in a ratio of 3:2:1 . Cryogenic Fourier transform infrared (FTIR) difference spectroscopy of the CO adduct of the reduced enzyme shows that the oxidase contains a heme-copper binuclear center and, thus, is a member of the heme-copper oxidase superfamily . In contrast to other members of this superfamily, however, this oxidase does not contain either heme O or heme A as a component of the binuclear center, but has heme B at this site . The single equivalent of Cu found in the oxidase is accounted for by the CuB component at the binuclear center . This suggests that this oxidase does not contain CuA, which is found in all other well-characterized cytochrome c oxidases . Both EPR and optical spectroscopic studies are consistent with this conclusion, also indicating that this oxidase does not contain CuA.(ABSTRACT TRUNCATED AT 250 WORDS) FEBS Lett, 1994 Mar 7, 340(3), 245 - 8 Modification of isolated subunit c of the F1Fo-ATPase from Propionigenium modestum by dicyclohexylcarbodiimide; Kluge C et al.; Subunit c of the F1Fo-ATPase from Propionigenium modestum was extracted from the particulate cell fraction with chloroform/methanol . The protein was further purified by carboxymethyl cellulose chromatography and anion exchange HPLC in the organic solvent . SDS-PAGE of the purified protein indicated a single stained protein band migrating as expected for the c-subunit . Incubation of isolated subunit c in chloroform/methanol or aqueous buffer containing dodecyl-beta-D-maltoside with {14C}dicyclohexylcarbodiimide (DCCD) resulted in the incorporation of radioactivity into the protein . The rate of this reaction depended on the external pH; it was significantly faster in the more acidic than in the alkaline pH range . In the presence of Na+ subunit c was partially protected from labeling with {14C}DCCD at pH 6.1 and at pH 7.5, whereas no protectio