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Volume 75 (1974)
- Issue 6 Pages 1201-
- Issue 5 Pages 951-
- Issue 4 Pages 679-
- Issue 3 Pages 441-
- Issue 2 Pages 215-
- Issue 1 Pages 1-
Volume 75, Issue 5
Displaying 1-31 of 31 articles from this issue
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Akira TAKETO1974 Volume 75 Issue 5 Pages 951-960
Published: May 25, 1974
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSSome properties of φA, the viral DNA and the intracellular RF were investigated. The virus particle is spherical and its diameter is about 300Å. φA is immunologically related to φX174 and physical properties such as optical rotatory dispersion were similar to those of φX174. The plating efficiency of φA on several E. coli strains was compared with those of other phages of φX174 group. The infectivity of the virion DNA is extremely sensitive to hydroxylamine treatment, UV* irradiation or DNase I [EC 3. 1. 4. 5], but is refractory to exonuclease I of E. coli. Spheroplast infectivity of φA DNA is inhibited by heterologous denatured DNA but not by native double-stranded DNA. Like φX174 DNA, φA DNA shows hyperchromicity upon treatment with formaldehyde. Its behaviour in methylated-albumin column chromatography or sucrose-density-gradient centrifugation of φA RF closely resembled that of φX174. The infectivity of φA RF increases several-fold on alkaline treatment, but is unaffected by incubation with hydroxylamine. Although φA RF is sensitive to DNase I, the inactivation rate is low as compared with the virion DNA. Survival of UV-irradiated RF is remarkably influenced by the uvr function of host cells and, in hcr+ bacteria, the UV sensitivity of RF is far lower than that of the virion DNA. The infectivity of φA RF to lysozyme-EDTA spheroplasts was inhibited specifically by double-stranded heterologous DNA, and infectivity of φA DNA to Ca2+-treated cells was inhibited by denatured DNA. These results show that while φA DNA is single-stranded, the RF molecule is double-stranded.View full abstractDownload PDF (1422K) -
I. Non-enzymatic Formation of Dipicolinic Acid from Pyruvate and Aspartic SemialdehydeKinuko KIMURA1974 Volume 75 Issue 5 Pages 961-967
Published: May 25, 1974
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSChemical condensation of pyruvate with aspartic, β-semialdehyde (ASA) results in the formation of two precursors of pyridine-2, 6-dicarboxylic acid (dipicolinic acid, DPA), with low and high mobilities on silica gel. Both act as substrates for dihydrodipicolinic acid reductase from Bacillus subtilis and also form adducts with o-aminobenzaldehyde. The reaction product of the pyruvate-ASA condensing enzyme from Bacillus subtilis seems to be identical with the DPA precursor with high mobility on silica gel. The non-enzymatic formation of DPA from the precursors is stimulated by Hg2+. A new method is described for the separation and estimation of DPA.View full abstractDownload PDF (534K) -
Susumu IWASA, Hitoshi AOSHIMA, Keitaro HIROMI, Hiroyuki HATANO1974 Volume 75 Issue 5 Pages 969-978
Published: May 25, 1974
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESS1) The Michaelis constant Km and molecular activity ko and/or the ratio ko/Km for linear substrates (degree of polymerization n ranging from 3 to 320) at 25°C and pH 5.85 were determined for two kinds of liquefying α-amylase [EC 3. 2. 1. 1] from two different strains of Bacillus subtilis. Little difference in n-dependence of the rate parameters was observed between the two amylases.
2) The number of effective subsites in the active sites of the enzymes and their affinities were evaluated from the experimental results. A procedure for evaluating subsite affinities from the n-dependence of ko/Km was proposed, which is applicable to the case where two or more productive complexes are involved. The values of subsite affinities of the two enzymes were found to be similar to each other. They were also similar to those obtained by Thoma et al. by the product analysis method for another liquefying α-amylase from a different strain of Bacillus subtilis.
3) Modes of cleavage of maltooligosaccharides observed qualitatively by thin layer
chromatography were essentially in agreement with those predicted from the evaluated subsite affinities.View full abstractDownload PDF (741K) -
IV. Hydrolyses of Synthetic Insulin Fragments by TrypsinNorikazu NISHINO, Nobuo MITSUYASU, Nobuo IZUMIYA1974 Volume 75 Issue 5 Pages 979-985
Published: May 25, 1974
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSA study was carried out to determine why the Arg-Gly bond in oxidized B-chain of insulin is hydrolyzed more rapidly by trypsin [EC 3. 4. 21. 4, formerly 3. 4. 4. 4] than the Lys-Ala bond in the same substrate. Three insulin fragments (Gly-Glu-Arg-Gly, Gly-Glu-Arg-Gly-Phe, and Thr-Pro-Lys-Ala) and a model peptide (Thr-Pro-Lys-Ala-Phe) were synthesized, and their susceptibilities to hydrolysis by trypsin were determined. No appreciable difference in susceptibility between the two tetrapeptides and the two pentapeptides, respectively, was observed. On the other hand, Gly-Glu-Arg-Gly-Phe was hydrolyzed 53 times faster than Thr-Pro-Lys-Ala. Thus, it is concluded that the difference in the lability of the two bonds in oxidized B-chain may be attributed to differences in the number of amino acid residues linked to the carboxyl groups of Arg and Lys residues.View full abstractDownload PDF (531K) -
Isamu SHIIO, Shinichi SUGIMOTO, Ryuichi MIYAJIMA1974 Volume 75 Issue 5 Pages 987-997
Published: May 25, 1974
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSStudies were made on the regulation and properties of 3-deoxy-D-arabino-heptulosonate-7-phosphate synthetase [7-phospho-2-keto-3-deoxy-D-arabino-heptonate D-erythrose-4-phosphate-lyase (pyruvate-phosphorylating), EC 4. 1. 2. 15] (DAHP synthetase*) of Brevibacterium flavum. The enzyme activity was stabilized by cobalt ion during purification and storage. Activity was maximal at pH 7.2 in Tris-HCl buffer. This enzyme required erythrose 4-phosphate and phosphoenolpyruvate as substrates, but at high concentrations they were inhibitory. The enzyme activity was inhibited by p-hydroxymercuribenzoate and this inhibition was overcome by adding dithiothreitol. Individually tyrosine or phenylalanine inhibited the reaction slightly, but together they caused strong, synergistic inhibition. Addition of 0.013mM tyrosine and phenylalanine at pH 7.5 caused 50% inhibition and the inhibition was maximal at pH 8.0. Other amino acids and intermediate metabolites, including tryptophan, p-aminobenzoate, chorismate and prephenate were not inhibitory. Among the phenylalanine analogues tested, m-fluorophenylalanine, β-thienylalanine and α-methylphenylalanine inhibited the activity only when tyrosine was present. DAHP synthetase of m-fluorophenylalanine resistant mutants which accumulated both tyrosine and phenylalanine in the culture medium, was not inhibited by tryosine plus phenylalanine. In the wild type strain of B. flavum, formation of DAHP synthetase was not affected by tyrosine, phenylalanine, and/or tryptophan, but in a phenylalanine auxotroph under phenylalanine-limiting condtions, over-production of tyrosine strongly repressed the enzyme, while in tyrosine auxotroph under tyrosine-limiting conditions, enzyme production was slightly derepressed. Tryptophan markedly reversed the repressive action of tyrosine. A mechanism for regulation of biosynthesis of aromatic amio acids in B. flavum is proposed on the basis of these results.View full abstractDownload PDF (777K) -
I. Further Purification and Some PropertiesJunichi SHIMIZU, Goro TAMURA1974 Volume 75 Issue 5 Pages 999-1005
Published: May 25, 1974
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSNitrite reductase [EC 1. 7. 7. 1] was purified approximately 1, 200-fold from spinach leaves, though the preparation was not homogeneous disc-electrophoretically. The purified enzyme catalyzed the reduction of nitrite, using reduced methyl viologen and spinach ferredoxin as an electron donor.
The enzyme showed three absorption peaks, at 278, 388, and 573mμ; the peak at 388mμ was shifted to longer wavelength and the peak at 573mμ disappeared upon reduction with sodium dithionite.
The apparent Michaelis constant of nitrite for the enzyme was estimated to be 5.55×1O-4M.
Although an electrophoretically homogeneous preparation could be obtained by preparative disc electrophoresis, the absorption spectrum of the enzyme was modified by this treatment, and the preparation showed methyl viologen-linked nitrite reductase activity but not ferredoxin-linked activity. The latter activity was restored by the addition of ferric ions. The molecular weight of this treated enzyme was estimated to be 63, 500 by SDS-polyacrylamide gel electrophoresis.View full abstractDownload PDF (1445K) -
Yasushi TOMITA, Akira OHASHI, Goro KIKUCHI1974 Volume 75 Issue 5 Pages 1007-1015
Published: May 25, 1974
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSThe induction of δ-aminolevulinate synthetase [EC 2. 3. 1. 37] by allylisopropylacetamide (AIA) and 3, 5-dicarbethoxy-1, 4-dihydrocollidine (DDC) was studied in a newly devised organ culture of finely chopped chick embryo liver. Both AIA and DDC appeared to stimulate the synthesis of ALA synthetase by acting primarily at the transcriptional level rather than the post-transcriptional level of protein synthesis. The half-life of ALA synthetase induced by AIA in our organ culture system was 2hr, whereas that of the enzyme induced by DDC was as long as 4-5hr. In the chick embryo liver, whether cultured or in ovo, accumulation of ALA synthetase in the cytosol fraction after induction by either AIA or DDC was very small, and the apparent molecular weight of the mitochondrial ALA synthetase from the chick embryo liver was about 250, 000 when estimated by gel filtration. Addition of hemin to the culture medium resulted in a considerable inhibition of ALA synthetase induction, probably by interfering mainly with a post-transcriptional spep(s). However, hemin did not cause appreciable accumulation of ALA synthetase in the liver cytosol fraction. Appararently the behaviour of ALA synthetase induction in the chick embryo liver differs in many respects from that in the adult chicken liver.View full abstractDownload PDF (734K) -
Kenji DOI, Akemi DOI1974 Volume 75 Issue 5 Pages 1017-1026
Published: May 25, 1974
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSA procedure was developed for isolating nuclei from cells of a tetraploid strain of Saccharomyces cerevisiae. In this procedure, spheroplasts, prepared from young yeast cells using the two bacterial glucanases, were lysed (a) at low osmotic pressure in the presence of a detergent or (b) at a moderate osmotic pressure in the absence of detergent, and the lyzate was homogenized and fractionated by centrifugation through discontinuous sucrose gradients. This procedure gave purified nuclei in 20-30% yield. The nuclei isolated using condition (a) for lysis were devoid of a nuclear envelope, while those isolated using condition (b) retained their envelopes. The nuclei isolated using either of the conditions of lysis had two to three times more RNA than DNA, and retained RNA polymerase [EC 2. 7. 7. 6] and NAD pyrophosphorylase [EC 2. 7. 7. 1] activities.View full abstractDownload PDF (3076K) -
I. Purification and Antiserum InhibitionHisayoshi TERAI1974 Volume 75 Issue 5 Pages 1027-1036
Published: May 25, 1974
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESS1. Adenylate kinase [EC 2. 7. 4. 3] has been purified approximately 200-fold from Pseudomonas denitrificans. The purified enzyme was nearly homogenous on polyacrylamide gel disc electrophoresis.
2. The molecular weight of the enzyme was estimated to be 21, 000 by the gelfiltration method. The heat stability, Km value and sensitivity to SH-inhibitors were compared with those of enzyme purified from other sources.
3. Antiserum against purified enzyme has been prepared from immunized rabbit. This antiserum inhibited 100 percent of the enzyme activity. Ouchterlony double diffusion tests showed one major and a few minor precipitin bands between the purified enzyme and the antiserum.
4. Immunological similarities of adenylate kinases from various sources were examined by comparing the degrees of inhibition of the activities by the antiserum.View full abstractDownload PDF (2198K) -
The Functional Role of Non-ionic Materials in Collagen Fibril FormationMasura HONYA1974 Volume 75 Issue 5 Pages 1037-1045
Published: May 25, 1974
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSIn order to investigate the role of non-ionic materials in an extracellular matrix in collagen fibrillogenesis, the effects of dextrans and polyoxyethylenenonyl phenyl ethers (poly-ONPE) on the process of collagen fibril formation were studied turbidimetrically and microscopically in an in vitro solute system at various concentrations.
1. In the presence of poly-ONPE's with hydrophilic polyoxyethylene chains of different lengths, -(CH2CH2-0-)n-, more hydrophilic poly-ONPE's markedly stimulated the rate of collagen fibril formation. The addition of dextrans of various molecular weights was also found to increase the rate of collagen fibril formation. Both accelerative effects were similar in pattern when they were plotted against the chain lengths.
2. In 0.1M urea-inhibited samples, poly-ONPE's with various hydrophilic chain lengths not only suppressed the inhibition by urea but, when highly hydrophilic, also accelerated the rate of collagen fibril formation.
3. Little change in viscosity was observed in collagen solutions containing poly-ONPE's with various hydrophilic-lipophilic balances.
4. On the basis of both microscopic and turbidimetrical observations, this accelerative effect was considered to result from interaction between the hydrophilic chains of the non-ionic materials and collagen molecules.
It seemed that the enhanced precipitation of collagen molecules resulted from the orderly alignment of water-bound non-ionic materials and the resulting “excluded volume” effect.View full abstractDownload PDF (764K) -
I. Purification and PropertiesKiyohisa MIZUMOTO, Kentaro IWASAKI, Michiko TANAKA, Yoshito KAZIRO1974 Volume 75 Issue 5 Pages 1047-1056
Published: May 25, 1974
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSThe eukaryotic polypeptide elongation factor 2 (EF-2) has been purified from pig liver homogenate by acid precipitation, calcium phosphate gel adsorption, ammonium sulfate fractionation, two successive DEAE-Sephadex column chromatographies and CM-Shephadex column chromatography. The purified EF-2 appears to be homogeneous as judged by gel electrophoresis and ultracentrifugation. Its specific activity is about 3, 000nmoles of phenylalanine polymerized in 30min at 37°C per mg of protein under standard assay conditions using poly (U), ribosomes from Artemia salina, and pig liver EF-1. It is stable at pH's between 8 and 10, and its stability increases markedly in the presence of 10mM 2-mercaptoethanol and even further in the presence of both 5mM magnesium ions and 0.2mM GTP or GDP. Some properties of polyphenylalanine synthesis reaction are also described. As with rat liver EF-2, the pig liver enzyme is inhibited by fusidic acid and also by diphtheria toxin and NAD+.View full abstractDownload PDF (1614K) -
II. Molecular Weight and Amino Acid CompositionKiyohisa MIZUMOTO, Kentaro IWASAKI, Yoshito KAZIRO, Chuuhei NOJIRI, Yu ...1974 Volume 75 Issue 5 Pages 1057-1062
Published: May 25, 1974
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSPolypeptide elongation factor 2 (EF-2) was purified from pig liver to a homogeneous state, as described in the preceding paper. In this paper we describe some of its molecular and physico-chemical properties, including molecular weight, sedimentation coefficient and amino acid composition.
The molecular weights determined by high-speed equilibrium centrifugation, SDS polyacrylamide gel electrophoresis and gel filtration on Sephadex G-200 were 100, 000, 95, 000, and 110, 000, respectively. The sedimentation coefficient at infinite dilution (s°20, w) was 5.32S.
The amino acid composition of pig liver EF-2 was also determined. Comparison of the results of amino acid analysis with those of EF-G from E. coli showed that there is a striking resemblance in amino acid composition, except that the halfcystine content in EF-2 is much higher than in EF-G.View full abstractDownload PDF (406K) -
Partial Purification and Some Properties of Pineapple CarboxypeptidaseEtsushiro DOI, Chizuko OHTSURU, Teruyoshi MATOBA1974 Volume 75 Issue 5 Pages 1063-1071
Published: May 25, 1974
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSSeveral commercial preparations of crude bromelain [EC 3. 4. 22. 4] from pineapple stem contained an additional proteolytic enzyme, carboxypeptidase (CPase). CPase in extracts from commercial bromelain powder was separated from bromelain by chromatography on CM-Sephadex C-50 at pH 4.5. Carboxypeptidase from pineapple stem was purified 300-fold from commercial bromelain powder by ion exchange chromatography and gel filtration. The enzymatic properties of pineapple CPase are as follows.
1. The enzyme is optimally active at pH 5.2 for carbobenzoxy-L-phenylalanyl-L-alanine (CPA) and is most stable around pH 5.2. The stability is increased by the addition of sucrose.
2. The enzyme activity is completely inhibited by diisopropyl fluorophosphate and partly inhibited by HgCl2.
3. The rate of hydrolysis of carbobenzoxy-L-glutamyl-L-tyrosine is about one-fifteenth of that for CPA.
4. The enzyme has esterolytic activity against acetyl-L-phenylalanine ethyl ester at neutral pH.
The similarity of these enzymatic properties of pineapple CPase to those of other plant CPases and pig kidney cathepsin A is discussed.View full abstractDownload PDF (689K) -
Tadao SAKAMOTO, Hiromichi OKUDA, Setsuro FUJII1974 Volume 75 Issue 5 Pages 1073-1079
Published: May 25, 1974
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSIt was demonstrated that rat liver homogenates contain high sterol-ester hydrolase activity. The activity was low in intact homogenates but increased considerably after treating the homogenates with Triton X-100. The bulk of the hydrolytic activity was associated with the microsomal fraction. The sterol-ester hydrolase [EC 3. 1. 1. 13] was found to be solubilized from the microsomal fraction after treatment with Triton X-100, suggesting that Triton X-100 activated the enzyme by solubilization.
The optimum pH of the enzyme was found to be around 6.7.
Among various cholesterol esters tested, cholesterol acetate was the most suitable substrate. Lysolecithin also activated the enzyme to the same extent as Triton X-100. Lecithin and deoxycholate failed to increase the activity of the enzyme. It was suggested that mechanism of activation by lysolecithin is different from that by Triton X-100.View full abstractDownload PDF (473K) -
Takashi YAMAMOTO, Yutaka ORII1974 Volume 75 Issue 5 Pages 1081-1089
Published: May 25, 1974
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSAn ordinary preparation of bovine cytochrome oxidase [EC 1. 9. 3. 1] at a purity of around lOnmoles heme a per mg protein was resolved into six polypeptide components of 40, 000, 25, 000, 19, 000, 14, 000, 10, 000 and 8, 000daltons by SDS* polyacrylamide gel electrophoresis, although the last component appeared to be heterogeneous, involving even smaller fragments. On the other hand, when “proteinase-treated cytochrome oxidase, ” which was derived from the ordinary preparation by limited proteolysis and retained the same enzymic activity as the starting material, was subjected to SDS gel electrophoresis, only two major components of 14, 000 and 11, 000 daltons were observed. Therefore, it is possible that these two polypeptides are functionally essential. However, from a comparison of their amino acid compositions, the 11, 000-dalton component is suggested to have been derived from the 14, 000-dalton component by partial proteolysis. Based on these findings, a model of cytochrome oxidase is presented.View full abstractDownload PDF (725K) -
Yukifumi NAGAI, Toichiro HOSOYA1974 Volume 75 Issue 5 Pages 1091-1101
Published: May 25, 1974
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSA procedure to isolate high purity Na+, K+-ATPase-associated plasma membrane in good yield from pig thyroid cells has been developed. The procedure has the advantage that a comparatively large amount of tissue can be processed in a relatively short time, yielding a plasma membrane fraction with high Na+, K+-ATPase activity. Fraction M+L, which was obtained from cytoplasmic extract by centrifugation at 250, 000g•-min and contained most of the Na+, K+-ATPase activity, was employed as a starting material for purification. The plasma membrane was purified from this fraction by two discontinuous density gradient centrifugations. By this method about 0.2mg membrane protein was recovered from 1g of wet thyroid slices and the membrane was purified 9-16 fold over Fraction M+L on the basis of Na+, K+-ATPase activity. By both morphological and enzymatic criteria, the purified membrane fraction appeared to be fairly free of contamination by other intracellular organelles. Several enzymes known to be present in plasma membrane of other tissues were purified along with Na+, K+-ATPase. The chemical composition of the membrane also agreed well with that of the plasma membrane of other tissues.View full abstractDownload PDF (1680K) -
Masako KIKUCHI, Koscak MARUYAMA, Haruhiko NODA1974 Volume 75 Issue 5 Pages 1103-1111
Published: May 25, 1974
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSF-Actin prepared by extraction from acetone powder of rabbit skeletal muscle was studied by analytical ultracentrifugation. F-Actin solution in O.1M KCl, pH 7.1, showed two schlieren peaks in the sedimentation pattern: a fast one of 85S and a slow one of 45S. The sedimentation patterns varied with the length of the extraction time of the acetone powder. Furthermore the concentration of the 45S component decreased with increasing sedimentation rate.
From measurements of the concentration of 45S F-actin during an ultracentrifuge run, we have suggested that a proteinaceous component in the 45S F-actin was dissociated during the run, and the helical fibrous structure of 45S F-actin then transforms into the more flexible structure of 85S actin.View full abstractDownload PDF (4107K) -
I. Purification and Function of Protein Components Required for the ReactionHideo KOCHI, Goro KIKUCHI1974 Volume 75 Issue 5 Pages 1113-1127
Published: May 25, 1974
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSFour protein components which participate in the reversible glycine cleavage reaction were isolated from extract of Arthrobacter globiformis grown in a glycine medium. Two of those protein components (a pyridoxal phosphate enzyme designated as P-protein and a heat-stable acidic protein designated as H-protein) were homogeneous on SDS-disc electrophoresis. The apparent molecular weights of H-protein and P-protein were 20, 000 and 270, 000, respectively, as estimated by Sephadex gel filtration.
Decarboxylation of the glycine carboxyl group absolutely required both P-protein and H-protein, and an H-protein-bound intermediate of glycine decarboxylation, which should be in the -CH2NH2 form, could be isolated. Incubation of the H-protein-bound intermediate with C02 and P-protein yielded glycine, and incubation with T-protein (a THF-requiring enzyme) and THF gave methylene-THF. Hydrogen atoms at the α-position of glycine were not labilized throughout the whole process from glycine to methylene-THF. H-Protein contained a functional disulfide group, and decarboxylation of glycine appeared to be coupled with reductive cleavage of the disulfide group in H-protein, followed by the formation of an S-C bond between the H-protein-thiol and -CH2NH2 moiety of glycine. In the reverse reaction of glycine decarboxylation, in turn, the dithiol group of H-protein seems to be oxidized to disulfide. A tentative scheme is presented to explain the whole process of reversible glycine cleavage.View full abstractDownload PDF (2078K) -
Ken-ichi NAKAMURA, Kazuhiko KONISHI1974 Volume 75 Issue 5 Pages 1129-1133
Published: May 25, 1974
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSBrain microsomes prepared with 10mM NaHCO3 showed ATP-dependent Ca2+ uptake but not Mg2+, Ca2+-dependent ATPase [EC 3. 6. 1. 3] activity. The incorporation of γ-32P Of ATP into microsomal lipid, which was located at the interface by extraction with chloroform-methanol, was observed only when the reaction conditions were appropriate for ATP-dependent Ca2+ uptake. Cysteine inhibited both 45Ca uptake and 32P incorporation. The 32P-incorporated lipid was identified as triphosphoinositide. These results lead to the conclusion that the ATP-dependent Ca2+ uptake of brain microsomes is due to the binding of Ca2+ to microsomal protein-bound triphosphoinositide, which is formed from ATP and diphosphoinositide by the action of diphosphoinositide kinase [EC 2. 7. 1. 68].View full abstractDownload PDF (352K) -
IV. Purification and Properties of ExonucleaseIkuo SUHARA1974 Volume 75 Issue 5 Pages 1135-1141
Published: May 25, 1974
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESS1. The exonuclease from Acrocylindrium sp. was purified about 200 fold by means of acetone precipitation, DEAE-Sephadex A-50 column chromatography, Sephadex G-100 gel filtration and hydroxyapatite column chromatography.
2. The hydroxyapatite fraction of the exonuclease was free from endonuclease, ribonuclease [ribonucleate guaninenucleotide-2'-transferase (cyclizing), EC 3. 1. 4. 8 or ribonucleate nucleotido-2'-transferase (cyclizing), EC 3. 1. 4. 22], phosphodiesterase [EC 3. 1. 4. 1] and phosphomonoesterase [EC 3. 1. 3. 1 or 3. 1. 3. 2].
3. The exonuclease was activated by Mg2+ and completely inactivated by heat treatment (60°C, 5min, at pH 7.5).
4. The optimum pH for the exonuclease activity ranged from 10 to 12.
5. The molecular weight of the exonuclease was estimated to be about 70, 000 by gel filtration.
6. The exonuclease hydrolyzed RNA, poly I, poly A, poly U, poly C, heat-denatured DNA, and oligonucleotides into 5'-mononucleotides.View full abstractDownload PDF (470K) -
Yoshihiko URATANI1974 Volume 75 Issue 5 Pages 1143-1151
Published: May 25, 1974
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSFor polymerization of Salmonella flagellin into flagellar filaments, a D2O medium containing 0.15M NaCl and 0.01M phosphate buffer (pD 7.4) is more favorable than an H20 medium containing 0.15M NaCl and 0.01M phosphate buffer (pH 7.0). Evidence for this was obtained from kinetic and equilibrium experiments carried out by viscosity measurements. Flagellin polymerization is known to consist of two steps, nucleation and elongation. It was found that when a D20 medium contained monomeric flagellin at concentrations higher than 1Omg/ml, nucleation occurred rapidly, while this rarely happened in an H2O medium. In addition, elongation of short polymers in the presence of a particular concentration of monomer became faster when D20 medium replaced H2O medium. Polymers (flagellar filaments) produced in the two kinds of media were compared by circular dichroism (CD) measurements and electron microscope observations: They were found to be indistinguishable. The overall monomer-polymer equilibrium shifts towards monomer on raising the temperature. It was shown that the equilibrium temperature for a mixture of given concentrations of monomer and polymer was raised by several degrees when the medium was changed from H2O to D2O. At 25°, monomers in H2O and D2O media had very similar CD profiles, indicating that their conformations were similar, if not completely identical. However, the conformation in D20 medium was shown to be more heat-stable than that in H20 medium. The experimental results support the current view that when proteins are transferred to D20 media, intra and intermolecular hydrophobic bonds and/or hydrogen bonds are strengthened.View full abstractDownload PDF (1796K) -
Reduction and Regeneration of Nitroxide Spin-labelsKiyofumi MARUYAMA, Shun-ichi OHNISHI1974 Volume 75 Issue 5 Pages 1153-1164
Published: May 25, 1974
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSThe photosynthetic bacterium Rhodospirillum rubrum and chromatophores taken from it have been studied with hydrophobic spin-labels. Stearic labels and lecithin lebels bind to the chromatophores and give ESR spectra consisting of two components, one corresponding to labels situated in the lipid bilayer portion of the membrane and the other due to labels directly or indirectly associated with the membrane proteins. The cytoplasmic fluid obtained by cell rupture shows a strong reducing activity in the dark for nitroxide spin-labels in aqueous solution as well as in chromatophore membranes. The reducing activity of the fluid treated in a variety of ways has been investigated. Both the proteins and some low molecular weight compounds in the fluid are necessary for the activity. NADH or NAD++malate can replace the low molecular compounds. The protein components catalyze the reduction of nitroxides by the low molecular compounds, probably NAD++malate, in the cytoplasmic fluid. The reduction of membrane-bound nitroxides results from some redox interaction of the reductase system with proteins in the membrane. The reduced nitroxides in chromatophore membranes are rapidly oxidized on irradiation at λ>660nm. The photo-oxidation probably occurs coupled with the electron transport system of the chromatophores. The stationary signal level of membrane-bound labels under illumination is governed largely by a balance between dark-reduction by the reductase system and photo-oxidation by chromatophores and reflects the redox potential of the photosynthetic system in R. rubrum cells.View full abstractDownload PDF (924K) -
Kiyofumi MARUYAMA, Shun-ichi OHNISHI1974 Volume 75 Issue 5 Pages 1165-1168
Published: May 25, 1974
Released on J-STAGE: November 18, 2008
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Shosuke TAKEMURA, Hiroshi KASAI, Miki GOTO1974 Volume 75 Issue 5 Pages 1169-1172
Published: May 25, 1974
Released on J-STAGE: November 18, 2008
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Tsuneo SATO, Takeru IIJIMA1974 Volume 75 Issue 5 Pages 1173-1176
Published: May 25, 1974
Released on J-STAGE: November 18, 2008
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Ichitomo MIWA, Jun OKUDA1974 Volume 75 Issue 5 Pages 1177-1179
Published: May 25, 1974
Released on J-STAGE: November 18, 2008
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Takao NAKAMURA, Yoshiki SUGITA, Yoshimasa YONEYAMA1974 Volume 75 Issue 5 Pages 1181-1182
Published: May 25, 1974
Released on J-STAGE: November 18, 2008
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Takashi OBINATA, Koui TAKAHASHI1974 Volume 75 Issue 5 Pages 1183-1186
Published: May 25, 1974
Released on J-STAGE: November 18, 2008
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Tadashi YOSHIDA, Sakiko TAKAHASHI, Goro KIKUCHI1974 Volume 75 Issue 5 Pages 1187-1191
Published: May 25, 1974
Released on J-STAGE: November 18, 2008
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Tomoh MASAKI, Tadatoshi KINOSHITA1974 Volume 75 Issue 5 Pages 1193-1195
Published: May 25, 1974
Released on J-STAGE: November 18, 2008
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Masato OHE, Hisayuki MATSUO, Fumio SAKIYAMA, Kozo NARITA1974 Volume 75 Issue 5 Pages 1197-1200
Published: May 25, 1974
Released on J-STAGE: November 18, 2008
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