|
|
|
|
|
| Biochim Biophys Acta, 1976 Feb 5, 418(3), 397 - 403 Phosphorylation of ribosomal proteins from eukaryotes in homologous and heterogous cell-free systems; Vassileva Grancharova T et al.; The phosphorylation of ribosomal proteins from eukaryotes in homologous and heterologous cell-free systems has been studied . The ribosomes and protein kinases from yeast (Saccharomyces cerevisiae, strain Bu), wheat (Triticum vulgare) and rabbit (Orystolagus cuniculus) have been used . It has been found that five ribosomal proteins incorporate gamma-32P from ATP during the incubation of wheat ribosomes with wheat protein kinase . When the phosphorylation of isolated wheat ribosomal proteins was examined more phosphoproteins were detected . These data confirm the suggestion that the ribosomal structure affects the phosphorylation . Probably some ribosomal proteins remain hidden for the action of protein kinase . The results from the crossed experiments show that there is no barrier for phosphorylation of yeast ribosomes with liver protein kinase, of wheat ribosomes with yeast and liver protein kinases and of liver ribosomes with yeast and plant protein kinases . The wheat protein kinase does not phosphorylate the yeast ribosomes under these experimental conditions . Some differences in the set of phosphoproteins obtained with various protein kinases have been detected . These data suggest that the ribosomal protein phosphorylation is not highly species specific although it is not universal. Eur J Biochem, 1976 Feb 2, 62(1), 211 - 5 New coenzymically-active soluble and insoluble macromolecular NAD+ derivatives; Zappelli P et al.; Reaction in dimethyl sulfoxide of nicotinamide 8-bromoadenine dinucleotide with the disodium salt of 3-mercaptopropionic acid afforded nicotinamide-8-(2-carboxyethylthio)adenine dinucleotide, a new NAD+ analogue functionalized at the adenine C-8 position by an omega-carboxylic side chain . Carbodimide coupling of the latter derivative to high-molecular-weight water-soluble (polyethyleneimine, polylysine) and insoluble (aminohexy)-Sepharose) polymers gave the corresponding macromolecular NAD+ analogues . These derivatives have been shown to be enzymically reducible . The polyethyleneimine analogue showed a substantial degree of efficiency relative to free NAD+ with yeast alcohol dehydrogenase (47%) but a considerably lower one with rabbit muscle lactate dehydrogenase (3%); the polylysine analogue showed a low degree of efficiency with both enzymes (5-6%). Arch Dis Child, 1976 Feb, 51(2), 91 - 9 Defective opsonization . A common immunity deficiency; Soothill JF et al.; Serum opsonization of yeasts for phagocytosis by normal polymorphonuclear leucocytes was defective in 11 of 43 children with unexplained frequent infections . The children had a range of infections, largely bacterial, and only 3 had diarrhoea and rash in infancy . A similar defect in at least 6 of the 9 mothers of these children (of either sex), with normal function in the fathers, suggests that the defect was primary and was transmitted by an unusual form of dominant inheritance . Four of 72 healthy adults and 1 of 11 children with unrelated disease showed similar defective function, but the incidence of the defect in the patients with frequent infection was significantly greater than this . The defective function can be corrected, in vitro and in vivo, by normal plasma at concentrations too low to be effective alone . This suggests that there is a defective factor rather than an inhibitor, and that different factors are limiting in normal and in defective plasma . Sera from affected members of the same family do not correct each other, but defective sera from different families usually do. J Biol Chem, 1976 Jan 25, 251(2), 270 - 6 Studies on cytochrome oxidase . Preliminary characterization of an enzyme containing only four subunits; Phan SH et al.; The preparation of a four-subunit enzyme from yeast, capable of catalyzing the oxidation of ferrocytochrome c, is described . It is derived from proteins containing seven or five subunits by means of recycling exclusion chromatography in the presence of 0.1% sodium dodecyl sulfate . Its catalytic properties are similar to those of the parent enzyme . Gel electrophoresis of this preparation in the presence of sodium dodecyl sulfate reveals three bands, migrating with RF values that correspond to molecular weights of 14.6, 12.3, and 10.6 X 10(3), with the largest exhibiting an apparent 2:1 stoichiometry relative to the other two . Visible spectra in the region of 390 to 630 nm do not show any detectable difference from that of the parent cytochrome oxidase, while its heme a and copper content are raised to values around 20 nmol or ng atoms/mg of protein, respectively, corresponding to minimal molecular weights of 50 X 10(3) . The molecular weight determined by physical means equals 107 X 10(3) . Thus the enzyme probably contains two copies of each subunit . After extensive dialysis to remove as much as possible of the sodium dodecyl sulfate used in its preparation, this enzyme remains in solution in phosphate buffer in the absence of any added detergent, while under similar conditions the seven-subunit complex precipitates completely . A similar preparation can also be obtained from beef heart . The significance of these findings is discussed with respect to the role of the large subunits in the function as well as the biogenesis of the mitochondrial cytochrome oxidase complex. Biochim Biophys Acta, 1976 Jan 20, 420(1), 81 - 6 Cross partition and determination of net charge of the isoenzymes of enolase; Blomquist G; Enolase from bakers' yeast was separated into three isoenzymes by countercurrent distribution . The isoenzymes were partitioned in aqueous polymer two-phase systems containing positively charged trimethylamino poly(ethylene glycol) or negatively charged poly(ethylene glycol) sulphonate . The plots of the partition coefficient of each isoenzyme versus pH in the two biphasic systems intersect at pH equal to the isoelectric point . From slopes of the plots, the net charge of the isoenzymes at pH 6.57 was determined to be +2, -3, and -8 respectively. Clin Chem, 1976 Jan, 22(1), 67 - 9 Improved microscale assay for purine phosphoribosyltransferase activities; Schmidt R et al.; We describe an improved and rapid filter-type assay for purine phosphoribosyltransferase activities . 14C-labeled purine bases are used as substrates, and the equipment includes a sampling manifold and glass-fiber filters coated with polyethyleneimine-cellulose . The method is especially suited for assay of a large number of samples, and may be useful for other enzyme activity measurements. Antonie Van Leeuwenhoek, 1976, 42(4), 493 - 502 Hereditary respiration deficiency in Saccharomycodes ludwigii; Nagai S et al.; Saccharomycodes ludwigii, supposed to be "petite-negative," gave rise to respiration-deficient mutants when acriflavine and ultraviolet irradiation, respectively, were applied to this yeast, strain IFO 1194 . The frequency of such mutants was very low as compared with that in Saccharomyces cervisiae and other "petite-positive" yeasts . Cytochrome composition was characterized by spectrophotometry at the temperature of liquid nitrogen . The respiratory mutants examined contained cytochrome c unaltered in quality and quantity . Cytochrome b was often present only in small amounts though never absent, while cytochrome a + a3 was either present or absent . The respiratory mutants could form zygotes after conjugation with a wild-type culture of opposite mating type (alpha vs . a) . The hybridization and segregation analysis of spore tetrads showed the inheritance of respiratory mutant character to be either Mendelian or non-Mendelian and similar to that of pet (nuclear) and rho- (cytoplasmic) mutants, respectively, in Saccharomyces cerevisiae. Nahrung, 1976, 20(6), 649 - 60 {Studies on lysine availability with Tetrahymena pyriformis as the test organism}; Meinl M et al.; A series of experiments was performed to verify the suitability of the protozoon Tetrahymena pyriformis (T.P.) for determining the lysine availability of feeds . The multiplication rate was estimated with the aid of organism counting, turbidimetry and reduction of triphenyltetrazolium chloride . The volume of T.P . was calculated on the basis of length and breadth measurements, Within higher rates of multiplication, there was connected a tendency towards larger organism volumes . A comparison of the growth tests showed that only turbidimetry is able to reflect, under certain conditions, the efficiency of the test organism in protein synthesis . The values for lysine availability varied widely according to the kind and pretreatment of the feedstuff, but did in no case attain those obtained with animal experiments . T.P . is, therefore, scarcely suited as a test organism for determining the protein quality and amino-acid availability of feedstuffs. Bioinorg Chem, 1976, 6(3), 229 - 32 Thallium antagonism toward potassium dependent systems; Siegel BZ et al.; When a number of Group 1A chlorides were added to yeast solutions which were inhibited by 20 mM Tl+, the greatest restorative effect was shown by KCl, with a lesser effect by NH4Cl . Sodium, rubidium and cesium chlorides had no significant effect on Tl+-inhibited CO2 production, and lithium chloride inhibited the system even further . Removal of potassium ions by dialysis reduced CO2 production by about 70% and the restorative effect of potassium was markedly reduced when this ion was added in the presence of thallium. Genetika, 1976, 12(8), 104 - 9 {Post-radiation recovery and the ploidy factor}; Kabakova NM et al.; Radiobiologicae effects on extensively homozygous Sacharomyces cerevisiae strains of different ploidy from haploid to hexaploid (developed by W . Laskovski) were studied . Radiation (gamma-rays of 60Co) inactivation studies showed a minimum of resistance of haploid strains, a maximum of resistance of diploid or triploid strains and a decrease of resistance with further increasing genome number . The explanation of such dependence of radiosensitivity on ploidy is usually due to the increase of dominant lethal damages and the corresponding decrease of recessive lethal damages with the increase of ploidy . All studied strains (except haploids) were capable of recovery of radiation damages after their storage in non-nutrient media during postradiation period . Since haploids are inactivated almost exclusively by recessive lethal damages, one may suppose that reversible part of radiation damages is due to dominant lethal damages . Then an irreversible part of radiation injury must decrease with the increase of ploidy . Indeed, in studied strains an irreversible component of radiation injury was significantly reduced with the increasing genome number . Any correlation of the probability of recovery from the primary damages with ploidy was not discovered. Cytobios, 1976, 16(62), 125 - 32 Nuclear envelope inclusions demonstrated by freeze-fracture; Severs NJ; Inclusions in the perinuclear space of the nuclear envelope of human diploid (MRC-5) fibroblasts, limpet (Patella vulgata) haemocytes, and yeast (Saccharomyces cerevisiae) as observed with the freeze-fracture technique are described . The significance of these inclusions is discussed and it is tentatively concluded that they represent vesicles engaged in transporting macromolecules between nucleus and cytoplasm . Although the inclusions were infrequently observed, their demonstration in mammalian, invertebrate and lower eukaryotic cell types raises the possibility that this form of nucleocytoplasmic exchange may potentially be adopted under appropriate circumstances by the eukaryotic cell in general. Ann Clin Res, 1976, 8 Suppl 17, 56 - 63 The regulation of heme biosynthesis; Poulson R; Changes in the levels of the heme biosynthetic enzymes were studied in cells and protoplasts of Saccharomyces cerevisiae during glucose derepression and respiratory adaptation . On aerobic or anaerobic glucose derepression or in the presence of 3', 5' cyclic AMP the levels of beta-aminolevulinic acid dehydratase, protoporphyrinogen oxidase and ferrochelatase increased whereas the levels of the enzymes catalysing the reactions from porphobilinogen to protoporphyrinogen IX remained constant . In contrast, the level of beta-aminolevulinic acid synthetase decreased during aerobic glucose derepression and in the presence of 3', 5' cyclic AMP, but it remained unchanged during anaerobic derepression . The heme content of the cells increased during aerobic glucose derepression but no increase was observed during anaerobic derepression of the cells . It is concluded that, in yeast, the inhibitory effect of anaerobiosis on heme synthesis is due, at least in part, to the requirement for oxygen in the oxidation of protoporphyrinogen IX, and that the effect of glucose on heme biosynthesis is mediated via glucose repression of protoporphyrinogen oxidase and possibly ferrochelatase. Biochimie, 1976, 58(1-2), 155 - 72 {Network of interactions between unlinked genes: synergistic and antagonistic regulation of iso-1-cytochrome c, iso-2-cytochrome c and cytochrome b2 synthesis}; Clavilier L et al.; Five chromosomal genes, CYPI to CYP5 involved in the regulation of the synthesis of iso-1-cytochrome c, iso-2-cytochrome c and cytochrome b2 are described . The function of these genes was studied either by varying the proportion of the mutated and wild type alleles in the cell vy varing the growth conditions, or else by transforming the mutants into sigma-cytoplasmic petites . We have shown a network of genetic interactions which regulate the synthesis of three structurally different proteins : iso-1-cytochrome c, iso-2-cytochrome c and cytochrome b2, by two unlinked genes : CYC1 and CYP1, one of which (CYC1) is the structural gene by iso-1-cytochrome c . Within this network the interactions are proportional to the gene dosage and are either antagonistic or synergistic depending on the allele combination and the protein studied . The mutated alleles cyp1 stimulate the synthesis of iso-2-cytochrome c, inhibit the synthesis of iso-1-cytochrome c, while the cytochrome b2 synthesis is also inhibited but by a combination of cyp1 mutated alleles CYC1 wild type allele . Other loci, CYP2, CYP3, CYP4 and CYP5 were also studied in various allelic combinations . They show some interactions between them or with CYC1 locus but these interactions are different and less pronounced than those involving loci CYP1 and CYC1. Microbios, 1976, 17(67), 17 - 21 Simple assay for the condensation component enzyme (beta-ketoacyl synthetase) of fatty acid synthetase; Brown OR et al.; A simple assay is described for estimating the activity of the condensation component enzyme (beta-ketoacyl synthetase) of the yeast fatty acids synthetase complex . The radioactivity liberated as 14CO2 from {1,3-14C}malonyl-CoA was trapped in phenethylamine and measured by liquid scintillation spectroscopy . Three enzyme-catalysed steps are involved: acetyl-CoA transacylase, malonyl-CoA transacylase and beta-ketoacyl synthetase; however, beta-ketoacyl synthetase is rate-limiting . beta-Ketoacyl synthetase activity was made independent of subsequent enzyme activities of the complex by excluding NADPH from the assay, thus blocking beta-ketoacyl reductase and preventing fatty acid synthesis . By this assay beta-ketoacyl synthetase activity was about 0.28 of the activity of the complex for fatty acid synthesis, compared with approximately 0.001 for published assays . Several pyridine nucleotides and derivatives were tested after it was discovered that NADH stimulated beta-ketoacyl synthetase activity to a greater extent than could be accounted for by its reactivity in providing a pathway from acetoacetyl-enzyme to fatty acid synthesis . Presumably, the release of acetoacetate from the central sulphydryl of the complex is the rate-limiting step in the assay procedure. J Biol Chem, 1975 Dec 25, 250(24), 9313 - 21 Sequence variability and structure of D-glyceraldehyde-3-phosphate dehydrogenase; Olsen KW et al.; The amino acid sequences of pig muscle and of yeast glyceraldehyde-3-phosphate dehydrogenase are compared with the three-dimensional structure of the lobster muscle enzyme . Residues in sheet and helical regions, on the exterior and interior, in subunit and domain interfaces, as well as residues in the active site have been examined for evolutionary conservation . The residues in the first (NAD binding) domain (1-147) are less conserved than residues in the second (catalytic) domain (148-334) probably because there are fewer internal residues and fewer residues involved in interactions between subunits . Residues in subunit interface are conserved to a significantly greater extent than others, and those involved in catalysis are conserved most of all . Patterns of residues in helices and sheets follow those found for other proteins. Biochemistry, 1975 Dec 16, 14(25), 5438 - 44 Inactivation of rabbit, pig, and carp adenylate kinases by N6-o- and p-fluorobenzoyladenosine 5'-triphosphates; Hampton A et al.; N6-O- and p-fluorobenzoyladenosine 5'-triphosphates (IIIc and IIc, respectively) have been synthesized as potential adenosine 5'-triphosphate (ATP) site-directed reagents for enzymes . IIc and IIIc were substrates of yeast hexokinase; neither they nor the corresponding ADP derivatives inactivated yeast hexokinase or rabbit pyruvate kinase . IIc rapidly inactivated rabbit and carp muscle adenylate kinases; the effect is probably ATP site directed because N6-benzoyl-ATP did not inactivate and was a substrate (Vmax = 28 and 10%, respectively, that of ATP), and because of ATP retarded the inactivation . The inactivations followed pseudo-firsr-order kinetics; in the presence of 2.64 mM ATP at 0 degrees the half-life of the rabbit kinase was 210 min with 50 muM IIc and the half-life of the carp kinase was 130 min with 100 muM IIc . Adenylate kinase of pig muscle was inactivated by IIc in a manner similar to the rabbit and carp enzymes except that the rate of inactivation exhibited an inflexion . IIIc inactivated rabbit, pig, and carp adenylate kinases by pseudo-first-order kinetics; the rate constants for inactivation at 0 degrees were 9.1 X 10(-3), 1.3 X 10(-3), and 1.9 X 10(-3) min-1 and the apparent dissociation constants (K) of the IIIc-enzyme complexes were 710, 970, and 720 muM, respectively . From the substrate properties of IIIc alone and in admixture with ATP its dissociation constants (Ki) from the ATP sites of the enzymes were found to be 500, 700, and 845 muM, respectively . The similarity between the K and Ki values, together with marked retardation of the inactivations by ATP, indicates that IIIc is an ATP-site-directed reagent for the three adenylate kinases. J Biol Chem, 1975 Dec 10, 250(23), 8986 - 9 Saccharopine dehydrogenase . Substrate inhibition studies; Fujioka M; In the direction of reductive condensation of alpha-ketoglutarate and lysine, saccharopine dehydrogenase (N6-(glutar-2-yl)-L-lysine:NAD oxidoreductase (lysine-forming) is inhibited by high concentrations of alpha-ketoglutarate and lysine, but not by NADH . NAD+ and saccharopine show no substrate inhibition in the reverse direction . Substrate inhibition by alpha-ketoglutarate and lysine is linear uncompetitive versus NADH . However, when the inhibition is examined with alpha-ketoglutarate or lysine as the variable substrate, the double reciprocal plots show a family of curved lines concave up . The curvature is more pronounced with increasing concentrations of the inhibitory substrate, suggesting an interaction of variable substrate with the enzyme form carrying the inhibitory substrate . These inhibition patterns, the lack of interaction of structural analogs of lysine such as ornithine and norleucine with the E-NAD+ complex (Fujioka M., and Nakatani, Y . (1972) Eur . J . Biochem . 25, 301-307), the identity of values of inhibition constants of alpha-ketoglutarate and lysine obtained with either one as the substrate inhibitor, and the substrate inhibition data in the presence of a reaction product, NAD+, are consistent with the mechanism that substrate inhibition results from the formation of a dead-end E-NAD+-alpha-ketoglutarate complex followed by the addition of lysine to this abortive complex. Biochemistry, 1975 Dec 2, 14(24), 5274 - 9 The influence of pH on the interaction of inhibitors with triosephosphate isomerase and determination of the pKa of the active-site carboxyl group; Hartman FC et al.; Ionization effects on the binding of the potential transition state analogues 2-phosphoglycolate and 2-phosphoglycolohydroxamate appear to be attributable to the changing state of ionization of the ligands themselves, therefore it is unnecessary to postulate the additional involvement of an ionizing residue at the active site of triosephosphate isomerase to explain the influence of changing pH on Ki in the neutral range . The binding of the competitive inhibitor inorganic sulfate is insensitive to changing pH in the neutral range . 3-Chloroacetol sulfate, synthesized as an active-site-specific reagent for triosephosphate isomerase, is used to provide an indication of the pKa of the essential carboxyl group of this enzyme . Previously described active-site-specific reagents for the isomerase were phosphate esters, and their changing state of ionization (accompanied by possible changes in their affinity for the active site) may have complicated earlier attempts to determine the pKa of the essential carboxyl group from the pH dependence of the rate of inactivation . Being a strong monoprotic acid, chloroacetol sulfate is better suited to the determination of the pKa of the carboxyl group . Chloroacetol sulfate inactivates triosephosphate isomerase by the selective esterification of the same carboxyl group as that which is esterified by the phosphate esters described earlier . From the pH dependence of the rate of inactivation of yeast triosephosphate isomerase, the apparent pKa of the active-site carboxyl group is estimated as 3.9 +/- 0.1. Am J Med, 1975 Dec, 59(6), 796 - 802 Hyperuricosuria and increased tubular secretion of urate in sickle cell anemia; Diamond HS et al.; Seven young adults with uric acid overproduction due to sickle cell anemia were normouricemic with a mean serum uric acid level of 4.9 mg/100 ml . Urate clearance was greater in these patients than in normal subjects or in patients with primary hyperuricemia due to uric acid overproduction . The increase in urate clearance was entirely accounted for by increased pyrazinamide suppressible urate clearance . Pyrazinamide administration abolished the uricosuric response to ribonucleic acid (RNA) feeding in these patients with sickle cell anemia, and maximal uricosuric response to the administration of probenecid was similar in the patients with sickle cell anemia and in normal subjects suggesting that reabsorption of both filtered and secreted urate was not impaired in sickle cell disease . Pyrazinamide suppressible urate clearance at maximal uricosuric response to probenecid was increased in patients with sickle cell disease suggesting increased tubular secretion of urate . This increase in urate secretion permits most young adults with urate overproduction due to sickle cell anemia to remain normouricemic and may account for the low frequency of secondary gout in this disease. J Biochem (Tokyo), 1975 Dec, 78(6), 1347 - 52 Application of the enzymic electric cell method to the activity assay of NAD-linked dehydrogenases; Nakano K et al.; An enzymic electric cell was constructed with a saturated calomel electrode (cathode) and an enzymic electrode (anode) which consisted of a glassy carbon electrode and a mixture containing an NAD-linked dehydrogenase, NAD+, Nomethylphenazonium methosulfate and a substrate, and the short-circuit current of the cell was measured... Can J Biochem, 1975 Dec, 53(12), 1278 - 81 Assembly of complex III into newly developing mitochondrial membranes; Aithal HN et al.; Yeast cells grown anaerobically in 0.02% linoleic acid were transferred to air in the presence of 0.02% elaidic acid . At varying times Arrhenius plots were made of QH2-cytochrome c reductase activities in isolated mitochondria . A transition temperature of 8.2 degrees C at 0.5 h was characteristic of linoleate; at 3 h the transition temperature was increased to 24 degrees C characteristic of elaidate . At early times the enzyme was associated with anaerobic promitochondrial membranes; at later states the newly synthesized enzyme was associated with newly developed elaidate membranes. J Biol Chem, 1975 Nov 25, 250(22), 8591 - 7 Structural analyses of mammalian ribosomal ribonucleic acid and its precursors . Nucleotide sequence of ribosomal 5.8 S ribonucleic acid; Nazar RN et al.; The nucleotide sequence of ribosomal 5.8 S RNA (also known as 7 S or 5.5 S rRNA) from Novikoff hepatoma ascites cells has been determined to be (see article) . Estimations of the secondary structure based upon maximized base pairing and the fragments of partial ribonuclease digestion indicate that there may be five base-paired regions in the molecule, three forming a folding of the termini and two forming secondary hairpin loops . The sequence of Novikoff hepatoma 5.8 S rRNA is about 75% homologous with that of yeast 5.8 S rRNA (Rubin, G.M . (1973) J . Biol . Chem . 248, 3860-3875) and similar models for secondary structure are proposed . Both models contain a very stable G-C rich hairpin loop (residues 116 to 138), a less stable A-U-rich hairpin loop (residues 64 to 91) and two symmetrical bulges (residues 15 to 25 and 40 to 44). Hoppe Seylers Z Physiol Chem, 1975 Nov, 356(11), 1693 - 701 Characteristics of DNA fractionated on benzoylated DEAE-cellulose; Pirro G et al.; Chromatography on BD-cellulose columns with a salt gradient and formamide separates cellular DNA into two fractions (fraction I eluted within the salt gradient, fraction II with formamide), the proportions of these two fractions (ca . 2:1) being similar for DNA from a number of eucaryotic organisms . Yeast DNA was chosen for a detailed study of the mode of fractionation . Several physicochemical parameters, binding to nitrocellulose filters, sensitivity towards nuclease S1, labelling properties in vivo, and hybridization properties of the two DNA fractions were compared . It was shown that both fractions are native DNA and that the fractionation does not depend on the size or the (G + C) content of the DNA . Fraction I DNA contains only a small portion of molecules having single-stranded ends . Fraction II DNA is a heterogeneous population, containing molecules with peculiar structural characteristics: (a) It contains DNA molecules with single-stranded ends and/or gaps sensitive to nuclease S1; labeling experiments suggested that these are molecules undergoing repair and replication . (b) Another portion of fraction II is molecules sensitive to nuclease S1 in regions which are not single-stranded . (c) A third portion is DNA which, after treatment with nuclease S1, is still strongly bound to the resin . Indications that the segregation may be due to the presence of specific DNA sequences comes from the above experiments and from the finding that fraction I DNA is enriched in ribosomal genes and fraction II DNA in tRNA genes. Z Naturforsch {C}, 1975 Nov-Dec, 30(6), 734 - 8 Interaction between dehydrogenases and a new NAD -isomer; Jeck R et al.; A new NAD -isomer was prepared, in which the D-ribose of the adenosine moiety was substituted by the enantiomeric L-ribose . As compared to nicotinamide-adenine-dinucleotide (NAD) and NADH the coenzyme isomer (D,L)-NAD and its dihydroform (D,L)-NADH are far less tightly bound to lactate dehydrogenase and alcohol dehydrogenase from horse liver . In the presence of the second substrate (D,L)-NAD and (D,L)-NADH act as hydrogen acceptor and hydrogen donator, respectively, with lactate dehydrogenase and alcohol dehydrogenases from horse liver and yeast . Compared to NAD and NADH the Michaelis constants are always increased, the catalytic constants (V/Et) were found to be decreased except for the dihydroform reacting with alcohol dehydrogenase from liver. Biofizika, 1975 Nov-Dec, 20(6), 1068 - 72 {Incorporation of low molecular SH-containing compounds in nitrosyl complexes of nonheme iron in cell-free and cell preparations}; Vanin AF et al.; It has been shown that the endogenic low molecular SH-containing compounds in non-cellular liver and yeast preparates include in nitrosyl non-haem iron complexes at basic pH and in result of addition of iron salts into these preparates . The content of nitrosyl non-haem complexes with SH-containing low molecular compounds is not more than 0,5-1% of the quantity of identical non-haem iron complexes with RS-groups of proteins . The low content of these complexes appear to be due to as the low content of low molecular compounds with ionized RS-groups as the competitive action and RS-groups of proteins for iron and NO. J Bacteriol, 1975 Nov, 124(2), 736 - 9 Characterization of 5.8S ribosomal ribonucleic acid in Neurospora crassa; Lucas MC et al.; Neurospora crassa ribosomes contain a species of ribonucleic acid (RNA) of molecular weight 54,000, similar to 5.8S ribosomal RNA previously described for other eukaryotic organisms . The 5.8S RNA from N . crassa was found to be released by heat treatment at 60 C from 25S ribosomal RNA but not from 18S ribosomal RNA . The base composition of N . crassa 5.8S RNA was similar to that of 5.8S RNA from Saccharomyces cerevisiae, but differed from animal 5.8S RNA . During the course of this study, it was discovered that N . crassa 25S ribosomal RNA had a number of internal cleavages that may exist in vivo. J Gen Microbiol, 1975 Nov, 91(1), 127 - 38 Mitochondrial structure studied by high voltage electron microscopy of thick sections of Candida utilis; Davidson MT et al.; Mitochondrial structure in yeast cells under various physiological conditions has been studied by high voltage electron microscopy of sections that are 0-5 to 2-0 mum thick . Such thick sections of the yeast Candida utilis had a small number of long, branched tubular mitochondria per cell . The mitochondria extended into cell buds and unseparated daughter cells . It was apparent from parallel studies with thin sections that most of the rounded mitochondrial profiles viewed in thin sections should not be interpreted as being numerous small individual mitochondria . Attempts to study thick sections of the yeasts Saccharomyces cerevisiae and Schizossaccharomyces pombe were frustrated by poor contrast. Biochim Biophys Acta, 1975 Oct 20, 405(2), 492 - 9 Pyruvate decarboxylase III . Specificity restrictions for thiamine pyrophosphate in the protein association step, sub-unit structure; Gounaris AD et al.; Pyruvate decarboxylase dissociates into sub-units of one half the molecular weight at alkaline pH . At the same conditions the cofactors thiamine pyrophosphate and Mg2+ are released and can be separated from the protein . Thiamine pyrophosphate is an obligatory cofactor for reconstitution to the oligomer {1} . In this study the effect of thiamine pyrophosphate derivatives (thiamine monophosphate, thiamine, and thiazole pyrophosphate) upon the reconstitution procedure was evaluated . The complete association of sub-units to form active oligomer was attained only when thiamine pyrophosphate was present . It is concluded that both the pyrimidine ring and the pyrophosphate group are required for productive co-enzyme binding and it is proposed that this interaction effects a conformational change which promotes protomer aggregation to form the enzymatically active holoenzyme . In addition data are presented which indicate that the monomer unit is 60 000 +/- 3000 daltons and that the N-terminal amino acid is histidine . Since the molecular weight of the active oligomer is 230 000 it is proposed that pyruvate decarboxylase is a tetramer comprised of four identical or nearly identical monomer units. Experientia, 1975 Oct 15, 31(10), 1147 - 9 Colloidal gold granules as markers for cell surface receptors in the scanning electron microscope; Horisberger M et al.; A rapid method has been developed to visualize cell surface receptors in the SEM . Thus mannan at the surface of Candida utilis cells was localized by stabilized colloidal gold granules coated with either anti-mannan antibodies or Con A. Biochim Biophys Acta, 1975 Oct 15, 407(3), 308 - 19 Binding of ethidium bromide to ribosomal RNA . Absorption, fluorescence, circular and electric dichroism study; Gatti C et al.; The interaction between ethidium bromide and ribosomal RNA has been studied by means of absorption, fluorescence, circular and electric dichroism measurements in the near ultraviolet and visible regions at low ionic strength (1 . 10(-3) and 6 . 10(-3) . The results have been interpreted on the basis of a model of interaction involving the intercalation of the phenanthridinium ring of the dye in the double-stranded regions of the RNA molecule, resulting in an increase of the dye-dye interactions as compared to DNA, and a stiffening of the intercalation regions. J Biol Chem, 1975 Oct 10, 250(19), 7739 - 46 Use of diazido ethidium bromide as a specific probe for mitochondrial functions; Bastos RN; The diazido derivative of ethidium bromide has been synthesized as a potential photoaffinity label and shown to be at least as effective as a mitochondrial mutagen as the parent compound, with a similar mode of action . Exposure of mitochondria of Saccharomyces cerevisiae to the compound, followed by ultraviolet-irradiation, which converts it to the highly reactive dinitrene, results in its specific binding to a single component which has been tentatively identified as the smallest polypeptide (subunit 9) of the membrane-bound ATPase . An analogus reaction is also obtained with the soluble, oligomycin-sensitive ATPase complex but not with the F1-ATPase itself . The reaction with the ATPase complex can also be monitored by fluorescence enhancement and by this attribute, as well as by other criteria, diazido-ethidium bromide, ethidium bromide itself, euflavine, N,N'-dicyclohexylcarbodiimide, 2,4-dinitrophenol, and 2-azido-4-nitrophenol all appear to compete for the same, lipophilic, binding site . A mitochondrial mutation (73/1) (see Flury, U., Feldman, F., and Mahler, H.R . (1974) J . Biol . Chem . 249, 6630-6637) produces a photoaffinity product with an altered electrophoretic mobility and molecular weight. J Antibiot (Tokyo), 1975 Oct, 28(10), 737 - 42 A glyoxalase I inhibitor of a new structural type produced by Streptomyces; Takeuchi T et al.; Many streptomyces strains produced an inhibitor of crude glyoxalase prepared from rat liver which did not inhibit glyoxalase I prepared from yeast . Another inhibitor, C11H14O6, which inhibited glyoxalases prepared from both rat liver and yeast was obtained from a cultured broth of Streptomyces griseosproeus and crystallized . Preincubation of this inhibitor with reuduced glutathione increased its inhibitory activity, which suggested its reaction with reduced glutathione . It showed a strong inhibition of growth of HeLa cells and inhibition of Ehrlich ascites carcinoma by daily injection . It also showed weak inhibition of the solid type of Ehrlich carcinoma and prolonged the survival period of mice inoculated with L-1210 cells. Mutat Res, 1975 Oct, 30(1), 43 - 54 Studies on the induction of mitotic gene conversion by ultraviolet irradiation . II . Action spectra; Ito T et al.; Action spectra for the induction of intragenic mitotic recombination (gene conversion) at the trp 5 locus by UV are presented for three cell stages (T0, T9 and T16) taken from synchronously growing cultures of Saccharomyces cerevisiae . The spectra over the range from 230 to 300 nm were taken mostly in 5-nm steps . The peak of action spectra was significantly shifted, regardless of the stage, toward the longer wavelengths as compared with that of the absorption spectrum of DNA (258 nm) or even that of thymine (265 nm) . In one extreme case (T16), the peak was shifted 17 nm from the absorption peak of DNA . Further, the spectrum changed its shape as the cell stage advanced from non-dividing (unbudded) (T0) to a dividing phase (T16) . Furthermore, the induction cross section decreased by a large factor (about 40), regardless of the wavelength, in going from T0 to T16 . From observations of the high photoreversibility of induced conversions, the major primary damage was thought to be pyrimidine dimers in the DNA . One plausible explanation, though not quite satisfactory from the quantitative viewpoint for these findings was that the increasing RNA during growth would screen the incident UV differentially with respect to the stage . If this explanation is correct, thymine dimers may still be considered, in spite of the shifts and deformations in the action spectra, as the major primary damage that triggers the long series of processes leading to gene conversion . Conventional methods for obtaining action spectra are discussed in comparison with the present method, which was based on sensitivity parameter a in the proposed dose (t)-frequency (f) relation, f = (at)alpha (alpha is the multiplicity parameter). Mutat Res, 1975 Oct, 30(1), 33 - 42 Studies on the induction of mitotic gene conversion by ultraviolet irradiation . I . Analysis of dose-frequency relationship; Ito T et al.; The UV (270-nm) dose-frequency relationship for the induction of intragenic mitotic recombination at trp 5 locus in Saccharomyces cerevisiae was non-linear . Two parameters, alpha and a, in the proposed equation for the non-linear relationship f = (at)alpha were determined so as to fit the experimental data by the method of least squares . The analysis was extended over 5 cell stages during synchronous growth . It was found that (1) parameter alpha changed from 2.02 for unbudded small cells to 1.09 for the stage where the cell had finished the division of the nucleus, and (2) parameter a changed correspondingly from 7.25-10(-4) to 0.180-10(-4) sec-1 during the same period . One interesting outcome in this analysis was the deduction of a dose-dependent nature of relative sensitivity with respect to the stage . The determination of these two parameters enabled us to calculate dose-effect relationships beyond the limits of experimental restrictions . Such an "imaginary" relationship, calculated at an extremely low dose, revealed the existence of maximal sensitivity around the DNA synthesis period . It was further shown that this maximum would easily be masked even in the moderate dose range . Thus, we conclude that the validity of single dose comparisons is diminished unless alpha is constant regardless of the cell stage . Some considerations on the proposed parameters have been made in relation to the mechanisms of the induction of gene conversion by UV. J Clin Invest, 1975 Oct, 56(4), 905 - 13 Hereditary deficiency of the seventh component of complement; Boyer JT et al.; Deficiency of the seventh component of complement has been found in the serum of a 42-yr-old Caucasian woman who has Raynaud's phenomenon, sclerodactyly, and telangiectasia . Partial deficiency was found in the serum of the patient's parents and children, indicating a pattern of inheritance of autosomal codominance . Transfusion experiments indicated that exogenous C7 had a 91-h halk-life in the patient . There was no evidence for C7 synthesis after transfusion . No C7 inhibitors were detected in the patient's serum . The patient's serum was found to support the activation of complement by both the classical and properdin pathways to the C7 stage . The addition of C7 to the patient's serum permitted it to support hemolytic reactions initiated by either pathway . No defects could be detected in plasma or whole blood coagulation . The patient's serum was deficient in opsonizing unsensitized yeast particles in serum and in the generation of chemotactic factor by antigen-antibody complexes and endotoxin . Both deficiencies were corrected by the addition of C7 . These observations suggest a key role for C7 for in vitro yeast phagocytosis and chemotaxis generation . However, the patient's lack of infections indicates a relatively minor role for C7 in human resistance to infection. Biochem J, 1975 Oct, 151(1), 37 - 45 The investigation of substrate-induced changes in subunit interactions in glyceraldehyde 3-phosphate dehydrogenases by measurement of the kinetics and thermodynamics of subunit exchange; Osborne HH et al.; An investigation was made of changes in subunit interactions in glyceraldehyde 3-phosphate dehydrogenase on binding NAD+, NADH and other substrates by using the previously developed method of measurement of rates and extent of subunit exchange between the rabbit enzyme (R4), yeast enzyme (Y4) and rabbit-yeast hybrid (R2Y2) {Osborne & Hollaway (1974) Biochem . J . 143, 651-662} . The free energy of activation for the conversion of tetramer into dimer for the rabbit enzyme (R4 leads to 2R2) is increased by at least 12kJ/mol in the presence of NAD+ . This increase is interpreted in terms of an NAD+-induced 'tightening' of the tetrameric structure probably involving increased interaction at the subunit interfaces across the QR plane of the molecule {see Buehner et al . (1974) J . Mol . Biol . 82, 563-585} . This tightening of the structure only occurs on binding the third NAD+ molecule to a given enzyme molecule . Conversely, binding of NADH causes a decrease in the free energy of activation for the R4 leads to 2R2 and Y4 leads to 2Y2 conversions by at least 10kJ/mol . This is interpreted as a NADH-induced 'loosening' of the structures arising from decreased interactions across the subunit interfaces involving the QR dissociation plane . In the presence of NADH the increase in the rate of subunit exchange is such that it is not possible to separate the hybrid from the other species if electrophoresis is carried out with NADH in the separation media . In the presence of a mixture of NADH and NAD+ the effect of NAD+ on subunit exchange is dominant . The results are discussed in terms of the known co-operativty between binding sites in glyceraldehyde 3-phosphate dehydrogenases. Arch Dermatol Res, 1975 Sep 12, 253(2), 113 - 8 Inhibition of glucose-6-phosphate dehydrogenase activity by betamethasone and three of its esters with dermatological importance; Raab WP et al.; Betamethasone, betamethasone-17-valerate, betamethasone-17-benzoate, and betamethasone-17,21-diproprionate were investigated for their inhbitory action on glucose-beta-phosphate dehydrogenase (G-6-PDH) activity (pure enzyme from yeast, enzyme from human skin homogenate) . Between these four compounds, marked differences were encountered which could not be attributed to the presence of an esterified or unesterified steroid . According to these data it does not seem to be justified to consider betamethasone esters simply as the transport forms of the topically inactive betamethasone but one must consider the betamethasone esters having biochemical actions of their own. J Biochem (Tokyo), 1975 Sep, 78(3), 617 - 26 Kinetic studies of carboxypeptidase Y . III . Action on ester, amide, and anilide substrates and the effects of some environmental factors; Bai Y et al.; Kinetic parameters of carboxypeptidase Y are given for the hydrolyses of ester, amide, and anilide substrates . The kcat/Km values were compatible with those of chymotrypsin {EC 3.4.21.1} with a few exceptions . One ionizable group with a pK of around 5.8 was suggested to be involved in the free enzyme in hydrolyzing all the substrates, including peptide substrates . In addition, hydroxylaminolysis and the kinetic isotope effects of deuterium oxide indicated, with some reservations, a reaction mechanism which proceeds via the formation of an acyl intermediate. Mol Cell Biochem, 1975 Aug 30, 8(2), 89 - 96 Studies of 3-aminopyridine adenine dinucleotide phosphate; Anderson BM et al.; 3-Aminopyridine adenine dinucleotide phosphate (AADP) was prepared from NADP and 3-amino-pyridine through the pig brain NADase-catalyzed pyridine base exchange reaction . The purified dinucleotide was chemically characterized and spectral properties of the compound were determined . The importance of the application of AADP in studies of NADP-requiring biochemical processes was indicated by the demonstration of AADP as an effective inhibitor of five NADP-requiring enzymes, by the demonstration of the fluorescence enhancement on the binding of AADP to yeast glucose-6-phosphate dehydrogenase when glucose-6-phosphate is present, and by the functioning of AADP as a fluorimetric substrate for snake venom nucleotide pyrophosphatase. Biochemistry, 1975 Aug 26, 14(17), 3908 - 12 Enolase catalyzed beta,gamma-alpha,beta isomerization of 2-phospho-3-butenoic acid to (Z)-phosphoenol-alpha-ketobutyrate; Appelbaum J et al.; 2-Phospho-3-butenoic acid was synthesized and found to be a substrate for both yeast and rabbit muscle enolase (EC 4.2.1.11) . Enolase catalyzes the isomerization of 2-phospho-3-butenoic acid to (Z)-phosphoenol-alpha-ketobutyrate, a beta,gamma-alpha,beta isomerization . Nuclear magnetic resonance studies on the product indicate only one isomer is formed . This reaction provides indirect evidence in further support of a carbanion intermediate for the enolase reaction . 2-Phospho-3-butenoic acid is also a good competitive inhibitor of both yeast and rabbit muscle pyruvate kinase (EC 2.7.1.40). Biochim Biophys Acta, 1975 Aug 26, 397(2), 277 - 87 Affinity chromatography of enzyme cofactors: the separation of NAD on immobilised dehydrogenase colums; Das K et al.; 1 . Alcohol dehydrogenase (EC 1.1.1.1.) has been immobilised to aminoethyl-cellulose by glutaraldehyde, to DEAE-cellulose by an s-triazine derivative and to agarose using CNBr . Lactate dehydrogenase has been immobilised to the latter two supports . 2 . Their use for affinity chromatography of NAD was compared and alcohol dehydrogenase immobilised to CNBr-activated agarose chosen for detailed study due to the efficient coupling of applied enzyme and the specific nature of binding . 3 . The efficiency of coupling of alcohol dehydrogenase dropped from 94.5 to 72.2% when the applied load was increased from 18 to 54 mg/g activated agarose . Activity relative to free enzyme fell from 21 to 11% . The binding of NAD was maximal between pH 5.5 and 6 . With the lowest loading of enzyme, NAD binding fell from 450 to 320 mug/g support when the linear flow rate was increased from 0.84 to 3.95 cm/min . 4 . NAD was completely separated from a mixture with ATP, ADP and AMP . Separation from NMN and hydrolysed RNA and DNA was evidently possible . Immobilised alcohol dehydrogenase used for 34 binding experiments over a period of weeks maintained 60% of its original enzyme activity . 5 . The method was applied to yeast NAD following mechanical disruption of yeast, clarification and either ultrafiltration or hollow-fibre dialysis to permit separate purification of macromolecules and nucleotides. Nucleic Acids Res, 1975 Aug 8, 2(8), 1237 - 60 A chemical approach to studies of the three-dimensional structure of tRNA - alkylation with a reagent covalently bound to a "peculiar site'; Grachev MA et al.; Yeast valine tRNA1 was chemically modified with chlorambucil N-hydroxysuccinimide ester . tthe reagent was attached covalently to the valine residue of valyl-tRNA1Val under the conditions which prevented tRNA from alkylation . Chlorambucilyl-valyl-tRNA1Val thus obtained was separated from excess reagent and incubated in an aqueous solution at neutral pH in the presence of Mg++ions . Highly efficient intramolecular self-alkylation of chlorambucilyl-valyl-tRNA1Val took place . The chlorambucil residue bound covalently to the amino group of the valine residue of tRNA1Val alkylates the 5'-terminal phosphate group of the molecule, and its 3'-terminal sequence -A-C-C-A. Nucleic Acids Res, 1975 Aug 8, 2(8), 1291 - 303 Drosophila chromatin: an immunological study; Roberts DB et al.; Antibodies were prepared against chromatin, various chromosomal protein preparations and against cytoplasm from Drosophila larvae . These antibodies were used to study the distribution of antigens in chromatin and chromosomal protein preparations on double diffusion plates . Antisera from all of the mammals tested precipitated both chromatin and DNA on double diffusion plates run in water . This non-specific precipitation was removed by washing in 0.06M NaCL. Cancer Res, 1975 Aug, 35(8), 2191 - 8 Binding of {3H}benzo(a)pyrene to natural and synthetic nucleic acids in a subcellular microsomal system; Pietropaolo C et al.; Several carcinogens are bound covalently to cellular nucleic acids . This is also the case with polycyclic hydrocarbon carcinogens, but their precise mechanism of in vivo activation to reactive forms and the structure(s) of the nucleic acid adducts are not known . This study demonstrates that in the presence of rat liver microsomes and reduced nicotinamide adenine dinucleotide phosphate there is covalent attachment of tritiated benzo(a)pyrene (BP) to transfer RNA, DNA, certain synthetic polyribonucleotides, and an RNA species endogenous to the microsomal fraction . Evidence has been obtained that the binding occurs mainly to guanine and, to a lesser extent, adenine residues and is not simple an artifact of tritium exchange . The microsomal-mediated binding of {3H}BP to nucleic acids requires reduced nicotinamide adenine dinucleotide phosphate and in inhibited by 7,8-benzoflavone, glutathione, and magnesium . It is enhanced somewhat by the addition of styrene oxide, cyclohexene oxide, and trichloropropylene oxide . These results provide the first evidence that: (a) the microsome-mediated binding of {3H}BP to nucleic acids is not just due to tritium exchange; (b) a derivative of the hydrocarbon is covalently bound to the nucleic acid, and not simply intercalated; (c) there is a preferential binding to guanine residues; and (d) in addition to binding to exogenous nucleic acids, {3H}BP is bound to an RNA species present in the microsomes . Our data are consistent with but do not prove that nucleic acid binding of this polycyclic hydrocarbon proceeds via an epoxide intermediate.
|
© 2005
Transgalactic Ltd (manufacturer of Bioscreen C software) |
Privacy Statement | P.O. Box
1393, 00101 Helsinki, Finland,
Last modified: May 25, 2005
| ||||||
