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Volume 86, Issue 1
Displaying 1-37 of 37 articles from this issue
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Effect of Proteolytic Modification on the Catalytic and Regulatory PropertiesIsamu KAMESHITA, Katsura IZUI, Hirohiko KATSUKI1979 Volume 86 Issue 1 Pages 1-10
Published: July 01, 1979
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSPhosphoenolpyruvate carboxylase from Escherichia coli W was treated with ten proteases, and the effects of the treatments on the enzyme activity and sensitivity to effectors were investigated. Proteases such as trypsin, α-chymotrypsin, papain, and subtilisin inactivated the enzyme, whereas elastase, carboxypeptidase Y and leucine aminopeptidase had no effect on the enzyme activity. Elastase and carboxypeptidase Y, however, inactivated the enzyme in the presence of 1M urea. Subtilisin and α-chymotrypsin caused not only inactivation of the enzyme but also a significant desensitization to the effectors. DL-Phospholactate, a potent competitive inhibitor, markedly protected the enzyme from inactivation by subtilisin but did not protect it from desensitization to the effectors. Acetyl-CoA, fructose 1, 6-bisphos-phate, and GTP-the allosteric activators-protected the enzyme from subtilisin inactivation, while laurate, the other allosteric activator, accelerated the inactivation. These activators did not protect the enzyme from desensitization to themselves. In contrast, modification with subtilisin in the presence of L-aspartate, the allosteric inhibitor, caused an apparent transient activation of the enzyme. The enzyme modified in the presence of L-aspartate retained its sensitivity to L-aspartate, but the sensitivities to the other effectors were reduced to about one-half their initial values. Based on these results, a possible mode of desensitization of the enzyme by subtilisin modification and the possible existence of a multiplicity of conformational states of the enzyme, induced upon binding with the various effectors, are discussed.View full abstractDownload PDF (777K) -
Shuzo OHYAMA, Koichi MIZUSAKI, Hideaki TSUNEMATSU, Satoru MAKISUM1979 Volume 86 Issue 1 Pages 11-16
Published: July 01, 1979
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSPhenylthiazolones (PTAs) of arginine and its homologs and analogs, homoarginine, norarginine (α-amino-γ-guanidinobutyric acid), canavanine, and γ-hydroxyarginine, were prepared. A steady-state kinetic analysis of the trypsin [EC 3.4.21.4]-catalyzed hydrolysis reactions was carried out and the kinetic parameters for these internal thioesters were compared with those for normal linear ester substrates. PTA-γ-hydroxyarginine was so labile that hydrolysis by the enzyme could not be followed. PTA-arginine has a specificity constant (kcat/Km) com-parable to that for the Nα-unblocked arginine ester substrate, though the value is about 0.1% of that for a specific ester substrate, Nα-tosylarginine methyl ester. PTA derivatives of cana-vanine and homoarginine were hydrolyzed with kcztlKm values of the same order of magnitude as that for PTA-arginine. However, PTA-norarginine was much less susceptible to tryptic hydrolysis than PTA-homoarginine, while the linear esters of norarginine are known to be more susceptible than those of homoarginine.View full abstractDownload PDF (465K) -
Isamu SHIIO, Shin-ichi SUGIMOTO1979 Volume 86 Issue 1 Pages 17-25
Published: July 01, 1979
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSChorismate mutase of Brevibacterium flavum, a common enzyme in phenylalanine and tyrosine biosynthesis, was separated into two different component, A and B, with molecular weights of 250, 000 and 25, 000, respectively, by ammonium sulfate fractionation or gel-filtration. Both components were essential for the enzymatic activity. In the presence of the reaction substrate, chorismate, the two components associated reversibly to give an active enzyme complex with a molecular weight of 320, 000. Binding sites of the feedback inhibitors, phenylalanine and tyrosine, on the enzyme were localized on component A as determined by hybridization experiments with the wild-type and mutant components. Tyrosine repressed the synthesis of component B much more strongly than that of component A, while phenyl-alanine did not show any significant repressive effect on either component. The wild-type strain No. 2247 had four times more component A than component B. Elution patterns in gel, DEAE-cellulose or hydroxyapatite column chromatography as well as the disc-gel electro-phoretic pattern of chorismate mutase component A and 3-deoxy-n-arabino-heptulosonate 7-phosphate (DAHP) synthetase activities completely overlapped, suggesting the presence of a bifunctional protein having the two activities. In accord with this suggestion, chorismate mutase as well as DAHP synthetase was insensitive to feedback inhibition by phenylalanine and tyrosine in all the 3-fluorophenylalanine-resistant mutants tested that excreted both phenyl-alanine and tyrosine. All the phenylalanine and tyrosine double auxotrophs defective in chorismate mutase lacked component B but not A.View full abstractDownload PDF (1391K) -
IV. Interaction of Maltose Receptor with a Membrane-Bound Chemosensing ComponentOsamu KOIWAI, Hiroshi HAYASHI1979 Volume 86 Issue 1 Pages 27-34
Published: July 01, 1979
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSHighly purified maltose receptor of Escherichia coli was bound to Sepharose 4B via a long spacer and affinity chromatography was performed to isolate the membrane-bound proteins having affinity for the maltose receptor. The experiments were carried out either in the pres-ence of maltose or in the absence of maltose and the proteins adsorbed on the matrix were identified by two-dimensional gel electrophoresis. The results showed that the maltose receptor interacted with the product of tar gene, one of the methyl-accepting chemotaxis proteins, only in the presence of maltose.View full abstractDownload PDF (2012K) -
Kazuko OHGI, Masachika IRIE1979 Volume 86 Issue 1 Pages 35-44
Published: July 01, 1979
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESS(1) RNase Ms was inactivated by iodoacetate. The inactivation was most rapid at pH 6.0, and was inhibited in the presence of a denaturant such as 8 M urea or 6 M guanidine-HCl.
(2) Competitive inhibitors protected RNase Ms from inactivation by iodoacetate; the effect was in the order 2', (3')-GTP>2', (3')-AMP, 2', (3')-UMP_??_2', (3')-CMP. The order is not consistent with that of the binding constants of the 4 nucleotides towards RNase Ms (A>C>G>U).
(3) RNase Ms was inactivated with the concomitant incorporation of one molar equivalent of carboxymethyl group. The following evidence indicated that the carboxymethyl group was incorporated into the carboxyl group of an aspartic acid or glutamic acid residue. (i) The carboxymethyl group incorporated into RNase Ms was liberated by treatment with 0.1 N NaOH or 1 M hydroxylamine. (ii) The amino acid composition of carboxymethylated RNase Ms (CM RNase Ms) after acid hydrolysis is similar to that of RNase Ms.
(4) 14C-Labeled CM RNase Ms was digested successively with alkaline protease and amino-peptidase M. The radioactive amino acid released was eluted just before aspartate on an amino acid analyzer. After hydrolysis with 6 N HCI, glutamic acid was produced exclusively from the radioactive amino acid. The specific radioactivity of this amino acid calculated from the radioactivity and glutamic acid formed was practically the same as that of CM RNase Ms. Thus, it was concluded that a carboxymethyl group was incorporated at the carboxyl group of a glutamic acid residue of RNase Ms.
(5) CM RNase Ms bound with 2'-AMP to the same extent as native RNase Ms, but bound to a lesser extent with 2', (3')-GMP.
(6) Although the conformation of CM RNase Ms as judged from the CD spectrum was practically the same as that of native RNase Ms, the reactivity of CM RNase Ms towards dinitrofluorobenzene was different from that of native RNase Ms, indicating some difference in the conformation.
(7) These results indicate that one glutamic acid residue is involved in the active site of RNase Ms.View full abstractDownload PDF (730K) -
Tatsuhiko OHE, Yasuto WATANABE1979 Volume 86 Issue 1 Pages 45-53
Published: July 01, 1979
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSXanthine dehydrogenase has been purified to a homogeneous state from cell-free extracts of a strain of Streptomyces. The enzyme has a molecular weight of 125, 000 and consists of two subunits with a molecular weight of 67, 000. The isoelectric point is at pH 4.4. The enzyme exhibits absorption maxima at 273, 355, and 457 nm and contains FAD, iron, and labile sulfide in a molar ratio of 1 : 7 : 1 per subunit. Little molybdenum could be detected. The enzyme is most active at pH 8.7 and at 40°C, and is stable between pH 7 and 12 (at 4°C for24 h) and below 55°C (at pH 9 for 10 min). The activity is stimulated by K+ at a concentra- tion of 50 mM or more and also by keeping the enzyme at pH 9 to 11. The activity is inhibited by cyanide, Tiron, and p-chloromercuribenzoate and by adenine and urate. Among the compounds tested, hypoxanthine, guanine, xanthine, 2-hydroxypurine, and 6, 8-dihydroxy-purine are oxidized at considerable rates; hypoxanthine is the best substrate. NAD+ is the prefered electron acceptor. Km values of the enzyme for hypoxanthine, guanine, xanthine, and NAD+ are 0.055, 0.015, 0.15, and 0.11 mM, respectively. Marked differences in the properties of this enzyme compared to others are the activity towards guanine, which has a higher affinity for the enzyme than hypoxanthine and xanthine, and a higher reactivity with hypoxanthine than xanthine. The organism has been identified as Streptomyces cyanogenus.View full abstractDownload PDF (1407K) -
I. Thermal Denaturation and Effects of SaltsMotohisa OOBATAKE, Sho TAKAHASHI, Tatsuo OOI1979 Volume 86 Issue 1 Pages 55-63
Published: July 01, 1979
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSThe thermal transition of RNase T1 was studied by two different methods; tryptophan residue fluorescence and circular dichroism. The fluorescence measurements provide information about the environment of the indole group and CD measurements on the gross conformation of the polypeptide chain. Both measurements at pH 5 gave the same transition temperature of 56°C and the same thermodynamic quantities, ΔHtr (=120kcal/mol) and ΔStr (=360eu/mol), for the transition from the native state to the thermally denatured state, indicating simultaneous melting of the whole molecule including the hydrophobic region where the tryptophan residue is buried. Stabilization by salts was observed in the pH range from 2 to 10, since the presence of 0.5 M NaCl caused an increase of about 5°C to 10°C in the transition temperature, depending on the pH. The fluorescence measurements on the RNase T1 com-plexed with 2'-GMP showed a transition with ΔHtr=167kcal/mol and ΔStr=497eu/mol at a transition temperature about 6°C higher than that for the free enzyme. The large value of ΔHtr for RNase T1 indicates the highly cooperative nature of the thermal transition; this value is much higher than those of other globular proteins. Analysis of the CD spectrum of thermally denatured RNase T1 suggests that the denatured state is not completely random but retains some ordered structures.View full abstractDownload PDF (653K) -
II. Salt-Induced RenaturationMotohisa OOBATAKE, Sho TAKAHASHI, Tatsuo OOI1979 Volume 86 Issue 1 Pages 65-70
Published: July 01, 1979
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSIn the presence of high concentrations of the monovalent salts, sodium chloride and potassium fluoride, disulfide-reduced RNase T1 having four cysteinyl residues intact regenerates the spectral properties characteristic of native RNase T1, i.e., the fluorescence spectrum of the aromatic side chains and the ultraviolet circular dichroism spectrum. The folding of the polypeptide chain proceeded without formation of disulfide bonds to yield an enzymatically active species havingan activity toward RNA equivalent to 25% of that of the native enzyme at the same salt concentration of 2M. Unfolding of RNase T1 by a denaturant, urea, was suppressed in the presence of salts, and the salt-induced chain folding was observed spectro-scopically even in 6.9M urea solution. The salts also induced the chain folding of disulfide-reduced and modified (carboxymethylated or carboxamidomethylated) RNase T1 into the native conformation, as indicated by its spectroscopic properties, but did not restore the enzymatic activity.View full abstractDownload PDF (464K) -
Yoshitaka GOTO, Jun SHIMIZU, Taro OKAZAKI, Ryoiti SHUKUYA1979 Volume 86 Issue 1 Pages 71-78
Published: July 01, 1979
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSCytosol PEP carboxykinase has been purified to electrophoretic homogeneity from bullfrog liver homogenate. The enzyme is a single polypeptide chain with a molecular weight of approximately 72, 000-75, 000. The purified enzyme catalyzed oxaloacetate decarboxylation (nucleoside triphosphate-supported), phosphoenolpyruvate carboxylation, and an exchange reaction between oxaloacetate and [14C]HCO3-in the presence of ITP or GTP. Manganese is absolutely required for the enzyme-catalyzed phosphoenolpyruvate carboxylation, whereas it can be replaced by Mg2+ for the oxaloacetate decarboxylation and the exchange reaction. The optimal pH of each reaction is dependent on the divalent metal ion used. The dependence of the enzyme activity on Mn2+ is markedly different in the phosphoenolpyruvate carboxylation and the oxaloacetate decarboxylation reactions.View full abstractDownload PDF (572K) -
Satoko ISEMURA, Eiichi SAITOH, Kazuo SANADA1979 Volume 86 Issue 1 Pages 79-86
Published: July 01, 1979
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSThree basic peptides with extremely high proline contents were isolated from human whole saliva. The amino acid sequences of two of these proline-rich peptides comprising 57 and 38 residues were determined by conventional methods. The sequence suggested that the smaller peptide was derived from the larger one and also revealed the occurrence of characteristic repeating units within the molecules. The present study is the first to describe this structural feature of proline-rich proteins or peptides.View full abstractDownload PDF (516K) -
Katsuko KAKINUMA, Teruhide YAMAGUCHI, Mizuho KANEDA, Kohkichi SHIMADA, ...1979 Volume 86 Issue 1 Pages 87-95
Published: July 01, 1979
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSFree H2O2 released from human blood leukocytes during phagocytosis into the extracellular medium was highly reactive with the ferric form of HRP, forming an enzyme-substrate complex which was identical to HRP-H2O2 compound II. The formation of HRP-H2O2 compound II was employed for assaying the rates of H2O2 release by leukocytes upon addition of bacteria or myristate. The treatment of normal human blood leukocytes with myristate resulted in a marked stimulation of H2O2 release compared to phagocytizing cells. The activity of H2O2 release in response to myristate was found to be deficient in the leukocytes of two patients with chronic granulomatous disease. This assay method with myristate supplementation is so sensitive and specific that it should be useful for the diagnosis of chronic granulomatous disease.View full abstractDownload PDF (671K) -
Masato OKADA, Seiichi KAWASHIMA, Kazutomo IMAHORI1979 Volume 86 Issue 1 Pages 97-104
Published: July 01, 1979
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSPutrescine oxidase [EC 1.4.3.4] of Micrococcus rubens oxidizes many kinds of synthetic polyamines: triamines (spermidine types), tetramines (spermine types), and N-substituted putrescines. Polyamines possessing terminal 4-aminobutylimino groups in their structures were more active as substrates. Putreanine was oxidized at a rate comparable to that of putrescine, and was converted to 1-pyrroline and β-alanine. Activities and Km values for polyamines were affected by the substituent attached to the 4-aminobutylimino group of the polyamine, and especially by its methylene chain length. It was also found that two types of oxidation occurred in the oxidation of polyamines by putrescine oxidase. When the moieties attached to the 4-aminobutylimino groups in polyamines were less hydrophobic, these polyamines were oxidized at the secondary amino groups to form 1-pyrroline. Polyamines which contained a hydrophobic substituent attached to the 4-aminobutylimino group were oxidized at the terminal primary amino group of the 4-aminobutylimino moiety to form ammonia. N, N'-Bis(4-aminobutyl)-1, 3-diaminopropane ([II, 4-3-4]) and N-(4-aminobutyl)-N'-(3-aminopropyl)-1, 3-diaminopropane ([II, 4-3-3]) were oxidized to form 1-pyrrolinium salt derivatives as a result of oxidation of the terminal primary amino groups. It was concluded that the essential structure for substrates of putrescine oxidase is a 4-aminobutylimino group (NH2(CH2)4NH-).View full abstractDownload PDF (553K) -
Tatsurokuro TOCHIKURA, Hirosuke FUKUDA, Mitsuaki MORIGUCHI1979 Volume 86 Issue 1 Pages 105-110
Published: July 01, 1979
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSThe purification and properties of NADPH-linked glyoxylate reductase [EC 1.1.1.79] from baker's yeast were studied. Two active fractions (peak I and peak II) were isolated by DEAE-cellulose column chromatography. The peak I fraction was purified to homogeneity by the criteria of disc gel electrophoresis and tentatively designated glyoxylate reductase I. Its molecular weight was calculated to be 31, 000 from gel filtration measurements. The enzyme reduced glyoxylate 7 times faster than hydroxypyruvate and was specific for NADPH. The enzyme showed optimum activity between pH 5.5 and 7.2. The Michaelis constants for glyoxylate and NADPH were found to be 13 mM and 4 μM, respectively. The enzymic activity was not significantly affected by anions, except for nitrate and iodide, which were inhibitory.View full abstractDownload PDF (1021K) -
Yoshifumi NINOMIYA, Yutaka NAGAI1979 Volume 86 Issue 1 Pages 111-119
Published: July 01, 1979
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSGlycosaminoglycan synthesis during cell growth has been studied in terms of unit cell numbers, using 16-day-old embryonic chick tendon cell cultures. Hyaluronic acid production was found to be inversely proportional to the cell density, while the levels of sulfated-glycosaminoglycan synthesis remained constant. On the other hand, hyaluronic acid production remained constant during cell proliferation, though chondroitin sulfate synthesis increased rapidly during an actively growing phase of the cultured cells, and dermatan sulfate and heparan sulfate syntheses increased gradually.View full abstractDownload PDF (1256K) -
Akira HACHIMORI, Yoko SHIROYA, Atsuko HIRATO, Tsuneo MIYAHARA, Tatsuya ...1979 Volume 86 Issue 1 Pages 121-130
Published: July 01, 1979
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSDivalent cations were shown to affect the structure and thermostability of thermophilic inorganic pyrophosphatase [pyrophosphate phosphohydrolase EC 3.6.1.1] purified from Bacillus stearothermaphilus and thermophilic bacterium PS-3. The properties of the enzymes from the two sources were found to be very similar. The enzymes were very unstable to heat in the absence of divalent cations, being inactivated gradually even at 40°C. However, they became stable to heat denaturation in the presence of Mg2+ between pH 7.8 and 9.0. Similar induced thermostability was detected when Mn2+, Co2+, Ca2+, Cd2+, and Zn2+ were added, though the latter three cations were not essential for enzyme activity. On adding divalent cations, the optical properties such as absorption spectra, fluorescence spectra, and circular dichroism (CD) were changed. Gel filtration and disc electrophoresis revealed that the molecular weight of both enzymes was 5.4×104 in Tris-SO4, buffer and 11×104 in Tris-HCI buffer, suggesting monomer-dimer transformation. In the presence of divalent cations in Tris-SO4 buffer, the enzymes dimerized; this was confirmed by sedimentation velocity measurements. The enzymes in Tris-HCI buffer did not show thermostability unless divalent cations were added. The results in the present study indicate that binding of divalent cations to each enzyme caused some conformational change in the vicinity of aromatic amino acid residues leading to dimerization of the enzyme molecule so that it became thermostable. It was also suggested that histidyl residues play an important role in the thermostability induced by divalent cations on the basis of the pH dependencies of thermostability and CD spectra.View full abstractDownload PDF (793K) -
Takashi OBINATA, Tomoh MASAKI, Hiromi TAKANO1979 Volume 86 Issue 1 Pages 131-137
Published: July 01, 1979
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSAntibodies were formed against the myosin light chains isolated from chicken fast skeletal, slow skeletal, and cardiac muscle and the antigenicities of the light chains were compared by double immunodiffusion and immunoelectrophoresis. It was shown that fast light chains are immunologically different from light chains of slow and cardiac myosin, while the slow and cardiac muscle light chains have similar immunological characteristics; that is, the light chains of apparent molecular weight about 27, 000 daltons in SDS-acrylamide gel electrophoresis of slow and cardiac muscle are immunologically indistinguishable, and the other light chains of apparent molecular weight about 19, 000 daltons of both muscles include a common antigenic site.View full abstractDownload PDF (3498K) -
Kosuke ICHIHARA, Ikuko YAMAKAWA, Emi KUSUNOSE, Masamichi KUSUNOSE1979 Volume 86 Issue 1 Pages 139-146
Published: July 01, 1979
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSThe microsomes from rabbit intestinal mucosa which had been washed quickly and thoroughly with phenylmethylsulfonyl fluoride were found to catalyze the hydroxylation of fatty acids in the presence of NADPH and molecular oxygen. Myristic and palmitic acids were converted to the corresponding ω- and (ω-1)-hydroxy fatty acids, whereas lauric acid was converted only to 12-hydroxylauric acid, and capric acid, to 9- and 10-hydroxycapric acids together with an unknown polar acid. Among these fatty acids, both myristic and lauric acids appeared to be the most efficient substrates. The inhibition of the hydroxylation by SKF 525-A and carbon monoxide suggested that the activity depended upon cytochrome P-450. The specific activity of the fatty acid hydroxylation was almost constant along the small intestine, while the aminopyrine N-demethylation activity and the cytochrome P-450 content were highest at the proximal end of the intestine and progressively declined toward the caudal end. The cytochrome P-450 was solubilized from the intestinal microsomes and purified by 6-amino-n-hexyl Sepharose 4B chromatography. The partially purified cytochrome P-450 was active in fatty acid hydroxylation in combination with intestinal NADPH-cytochrome c reductase and phosphatidylcholine.View full abstractDownload PDF (593K) -
Masashi KOSAKAI, Zensaku YOSIZAWA1979 Volume 86 Issue 1 Pages 147-153
Published: July 01, 1979
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSThe stability of ester sulfates in porcine intestinal heparin to solvolytic desulfation (100°C, 9 h) with dimethylsulfoxide containing 2% pyridine was examined, in comparison with the case of dilute acid treatment (0.1M HCl, 100°C, 70min). The resulting heparin modifications were deaminated and the deamination products were fractionated by the procedures reported previously ((1978) J. Biochem. 83, 1567-1575). The yields of disulfated disaccharide (b-2) and monosulfated disaccharides (e-2-1 and e-2-3) indicated that 2-O-sulfates in L-iduronic acid residues of heparin were more labile than 6-O-sulfates in glucosamine residues to the dilute acid treatment, whereas the opposite was the case for the solvolysis. The product of heparin modification by solvolysis was similar to whale heparin in the distribution of ester sulfates.View full abstractDownload PDF (495K) -
Haruo TANAKA, Shoji SHIMIZU, Ruiko OIWA, Yuzuru IWAI, Satoshi OMURA1979 Volume 86 Issue 1 Pages 155-159
Published: July 01, 1979
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSThe inhibition of growth and cell wall synthesis by 3-amino-3-deoxy-D-glucose (3-AG), which is known to be one of the constituents of the kanamycin molecule and a metabolite of Bacillus sp., was almost completely overcome by glucosamine and N-acetylglucosamine in Staphylococcus aureus but scarcely affected by D-glucose and D-fructose. The antibiotic did not inhibit the incorporation of [14C]glucosamine and [3H]N-acetylglucosamine into the acidinsoluble fraction, but rather enhanced the incorporation of [14C]glucosamine. On the other hand, it inhibited the incorporation of D-[14C]fructose into the cell wall fraction but hardly affected the incorporation of D-[14C]fructose into the acid-insoluble fraction in the presence of penicillin G.
Based on these results, it is suggested that the site of primary action of 3-AG is the forma-tion of glucosamine-6-phosphate from D-fructose-6-phosphate, which is catalyzed by glucosamine synthetase [EC 2.6.1.16].View full abstractDownload PDF (326K) -
Masataka ISHINAGA, Ryuhei KANAMOTO, Makoto KITO1979 Volume 86 Issue 1 Pages 161-165
Published: July 01, 1979
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSThe outer membrane of Escherichia coli K-12 contained a smaller proportion of phospholipid molecular species with two unsaturated fatty acyl chains than did the cytoplasmic membrane. Proportions of phospholipid molecular species in the outer and cytoplasmic membranes changed in response to temperature changes. As the temperature increased, the content of 1-palmitoy1-2-cis-9, 10-methylenehexadecanoyl species increased. Translocation of phos-pholipids from the cytoplasmic membrane to the outer membrane and synthesis of various molecular species were observed.View full abstractDownload PDF (325K) -
Kunio MATSUI, Natsuyo JURI, Yoko KUBO, Sabu KASAI1979 Volume 86 Issue 1 Pages 167-175
Published: July 01, 1979
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSA synthesis of roseoflavin by Streptomyces davawensis from guanine through riboflavin was demonstrated. The lines of evidence are (1) incorporations of 14C of [2- and U-14C] guanine and [2-14C] riboflavin into roseoflavin, (2) no incorporation of 14C of [8-14C] guanine into roseoflavin, (3) localizations of 14C in roseoflavin, and (4) a decrease of specific radioactivity of roseoflavin formed from [2-14C] guanine on addition of riboflavin to the culture. The 14C atoms in roseoflavin formed were localized by radioactivity analysis of the NaOH-hydrolysis products, i. e., urea and 1, 2-dihydro-6-methy1-7-dimethylamino-2-keto-1-D-ribityl-3-quinox-alinecarboxylic acid (QC), a new substance. These hydrolysis products were identified by the isolation of dixanthylurea, decomposition with urease, and from the properties of QC and QC tetraacetate isolated. These findings suggest that the pyrimidine ring of guanine is conserved in the formation of roseoflavin from guanine through riboflavin.View full abstractDownload PDF (2222K) -
Masanobu JANADO, Yuki YANO, Hiroko NAKAMORI, Toshiro NISHIDA1979 Volume 86 Issue 1 Pages 177-182
Published: July 01, 1979
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSThe oligomers of bovine serum albumin were produced by controlled reaction with peroxidizing linoleic acid to examine their possible utility as calibration proteins in sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The polymerization was effected in reaction mixtures containing linoleic acid undergoing peroxidation in the presence of ascorbic acid, and conditions that yield soluble oligomers with a wide molecular weight distribution were established. The interaction of these soluble oligomers with sodium dodecyl sulfate exhibited a binding isotherm indistinguishable from that obtained with bovine serum albumin. Furthermore, sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the albumin oligomers conformed to the empirical relation of molecular weight to mobility that pertains to the use of these oligomers as standard molecular weight markers.View full abstractDownload PDF (2462K) -
Shuichi FURUTA1979 Volume 86 Issue 1 Pages 183-190
Published: July 01, 1979
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSThe pigeon breast muscle pyruvate dehydrogenase complex was resolved into three component enzymes: lipoate acetyltransferase, pyruvate dehydrogenase, and lipoamide dehydrogenase. The antibodies against each component enzyme were prepared. All of the antibodies against component enzymes precipitated the pyruvate dehydrogenase complex.
The enzyme complex was recovered as the immunoprecipitate from the extract of breast muscle of a pigeon that had received a single injection of L-[4, 5-3H]leucine. The immunoprecipitate was separated into each component enzyme by SDS-polyacrylamide gel electrophoresis. The relative isotopic leucine incorporations per mg of protein into each component enzyme 4 h after the injection were 10 : 0.9 : 1.4 : 2.7 for lipoate acetyltransferase, α- and β-subunit of pyruvate dehydrogenase, and lipoamide dehydrogenase, respectively. The half-lives of lipoate acetyltransferase, α- and β-subunit of pyruvate dehydrogenase, and lipoamide dehydrogenase were 7.7, 2.5, 2.6, and 1.8 days, respectively. These results indicate that the component enzymes of the pyruvate dehydrogenase complex were synthesized and degraded at different rates.View full abstractDownload PDF (1929K) -
Takaaki KAMEJI, Yasuko MURAKAMI, Shin-ichi HAYASHI1979 Volume 86 Issue 1 Pages 191-197
Published: July 01, 1979
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSThe effects of two putrescine analogs were studied on hepatic polyamine synthesis and cell proliferation, both of which were stimulated by food intake. Trans-1, 4-diamino-2-butene (diaminobutene), which is a potent competitive inhibitor of ornithine decarboxylase [EC 4.1.1.17] (ODC), repressed the induction of ODC and effectively inhibited the accumulation of putrescine in rat liver which was induced by the feeding of dietary protein. Unexpectedly, diaminobutene did not suppress DNA synthesis and mitotic activity in rat liver, suggesting that it can mimic the role of putrescine in cell proliferation. 1, 3-Diaminopropane effectively repressed the induction of ODC caused by food intake and also suppressed DNA synthesis and mitotic activity without affecting the accumulation of RNA or protein. The suppression of mitoticactivity by 1, 3-diaminopropane was reversed by a single injection of putrescine, spermidine, spermine, or diaminobutene. It was concluded that rapid accumulation of polyamines, especially putrescine, was a prerequisite for the later enhancement of DNA synthesis and cell proliferation in rat liver caused by food intake.View full abstractDownload PDF (549K) -
II. Properties and Distribution of Thyroglobulin Labeled In Vitro or In Vivo with Radioiodine, 3H-Tyrosine, or 3H-Galactose in Rat Thyroid GlandsShigeru MATSUKAWA, Toichiro HOSOYA1979 Volume 86 Issue 1 Pages 199-212
Published: July 01, 1979
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSWith the aim of obtaining information on the process of iodination of thyroglobulin, the properties and subcellular distribution of thyroglobulin labeled with radioiodine, 3H-tyrosine, or 3H-galactose were studied. The following results were obtained for 17-19S thyroglobulin isolated from rat thyroid lobes labeled in vitro. (a) The effect of sodium dodecyl sulfate (SDS) concentration (0.1-2.0 mM) on the dissociability of the proteins into 12S subunits showed that 3H-labeled, 131I-labeled, and preformed thyroglobulin behaved very differently; their dis-sociability decreased in that order. In addition, 0.3 mM SDS is most suitable for discriminating among these species. (b) The amount of 0.3mm SDS-resistant 131I-thyroglobulin increased with the time of incubation of the lobes or with the amount of iodine atoms incorporated by chemical iodination. (c) Digestion of 3H-tyrosine-labeled thyroglobulin showed that 3H-monoiodotyrosine and 3H-diiodotyrosine were present after incubation of the lobes for 180 min. (d) The dissociability of 3H-galactose-labeled 17-19S thyroglobulin was higher than that of 1311I-labeled protein, but its elution pattern on DEAE-cellulose chromatography resembled that of the latter. (e) 131I-Thyroglobulin was scarcely found in the incubation medium, although a considerable amount of 19S thyroglobulin was released into the medium during the incubation. As for the lobes, a significant amount of 131I-radioactivity as well as 3H-radioactivity was found in cytoplasmic particulates, especially in fractions containing apical vesicles and rough microsomes. On the other hand, the following results were obtained for 17-19S thyroglobulin isolated from rats injected with 125I. (a) Dissociability of the protein by 0.3 mM SDS and analysis of 125I-iodoarnino acids of pronase digests showed that the iodina-tion process was essentially similar to the case of in vitro incorporation, but was faster. (b) The effect of cycloheximide treatment showed that the relative reduction of 0.3 mM SDS-View full abstractDownload PDF (1071K) -
Toshihiro SUGIYAMA, Retsu MIURA, Toshio YAMANO1979 Volume 86 Issue 1 Pages 213-223
Published: July 01, 1979
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSThe reaction processes of adrenodoxin reductase with NADPH and NADH were investigated. The appearance of a new intermediate with a broad absorption band at around 520 nm has been detected by rapid-scan stopped-flow spectrophotometry. Although the formation of this intermediate is more rapid with NADPH than with NADH, the rates of the subsequent decay to the fully reduced state are almost identical (kobs values were 20.5 and 16.0s-1) These results indicate that the new intermediate is the complex formed between the oxidized enzyme and reduced pyridine nucleotide (enzyme-substrate complex), and that subsequent decay of the intermediate is caused by a two-electron transfer process from the reduced pyridine nucle-otide to the enzyme flavin.
On the other hand, spectral and kinetic properties in the steady state of the reoxidation reaction of the enzyme reduced with NADPH and with NADH were somewhat different. The rate of reoxidation of the enzyme under aerobic conditions from the reduced state to the oxidized state was 6.5 times faster when a 10-fold molar excess of NADH was used than when NADPH of the same concentration was used. This result is consistent with the fact that the NADH-dependent oxidase activity was 6.4 times greater than that dependent on NADPH. During reoxidation of the reduced enzyme under aerobic conditions in the presence of an excess of NADPH or NADH, the EPR spectra indicated the formation of the flavin semi-quinone radical species. Similarly, the formation of semiquinone was observed in the absorption spectrum with either NADPH or NADH under the same conditions as in the EPR mea-surement. The intensity of the semiquinone signal on EPR was considerably smaller with NADH than with NADPH. These results suggest that NADP+ complexed with the enzyme semiquinone protects the radical from oxidation by oxygen to a greater extent than NAD+, and consequently the semiquinone is easier to detect with NADPH than with NADH.View full abstractDownload PDF (836K) -
Tomofusa TSUCHIYA, Keiko TAKEDA1979 Volume 86 Issue 1 Pages 225-230
Published: July 01, 1979
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSExtrusion of sodium ion from cells of Escherichia coli was measured using an Na+ electrode. When oxygen was supplied to an anaerobic cell suspension, extrusion of Na+ was observed. The addition of glucose under anaerobic conditions also caused Na+ efflux. The extrusion of Na+ energized by respiration and glycolysis was completely inhibited by a proton conductor, carbonyl cyanide m-chlorophenylhydrazone. These observations are consistent with the view that Na+ transport occurs secondarily to H+ circulation. Interestingly, induction of the melibiose transport system, which is coupled to Na+, greatly enhanced Na+ transport activity.View full abstractDownload PDF (373K) -
Sadaichi TAKAGI, Tomohiko MORI1979 Volume 86 Issue 1 Pages 231-238
Published: July 01, 1979
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSPoly(A)-containing RNA from soybean seeds was isolated by affinity chromatography on poly(U)-Sepharose. Translation of poly(A)-containing RNA was inhibited by the addition of m7GMP. Treatment of the poly(A)-containing RNA with periodate followed by aniline (a chemical removal of 5'-terminal m7G) resulted in a marked reduction of mRNA activity. mRNA activity of the treated RNA was partially restored when S-adenosyl methionine (SAM) was included in the wheat germ cell-free system.
End groups of the poly(A)-containing RNA were labeled by oxidation with periodate and reduction with [3H]borohydride. Analysis of nuclease digests of 3H-labeled poly(A)-containing RNA showed the existence of materials with a charge between -2 and -3 and/ or between -3 and -4 which contain pyrophosphate linkages. These results imply the presence of 5'-terminal capped structures, m7G(5')ppp(5')N and m7G(5')ppp(5')Nm, in soybean poly(A)-containing RNA.View full abstractDownload PDF (599K) -
Michinori MIYAHARA, Haruhiko NODA1979 Volume 86 Issue 1 Pages 239-248
Published: July 01, 1979
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSThe electric dipole moment of rabbit skeletal myosin was estimated from the electric and flow birefringence properties. Myosin formed small polydisperse aggregates (0.2-1.1μM in length) with an apparent electric dipole moment of 5, 000-20, 000 Debye in aqueous urea or sodium pyrophosphate at low ionic strength. Permanent dipole moment contributed substantially to the apparent dipole moment. An anti-parallel association of myosin was suggested from the dependence of the apparent dipole moment on myosin concentration. Some interactions between myosin and C-protein were detected in 1M urea by flow birefringence and analytical ultracentrifugation studies. The apparent dipole moment of myosin aggregates was less dependent on myosin concentration in the presence of C-protein.View full abstractDownload PDF (1336K) -
Its Kinetic Properties and a Reconstitution of Gluconate OxidaseKazunobu MATSUSHITA, Emiko SHINAGAWA, Osao ADACHI, Minoru AMEYAMA1979 Volume 86 Issue 1 Pages 249-256
Published: July 01, 1979
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSD-Gluconate dehydrogenase solubilized and purified from the membrane of Pseudomonas aeruginosa showed a Km value that varied with pH, namely 0.8mM at pH 6.0 and 3.2mM at pH 5.0. The enzyme was highly specific for D-gluconate and had an optimum pH 5.0-6.0. These kinetic properties of the solubilized enzyme were basically the same as those of the native enzyme in the membrane. The enzyme activity was inhibited competitively by pyruvate and 2-ketogluconate, non-competitively by oxamate and in a mixed-type fashion by oxalate.
The enzyme could reduce phenazine methosulfate, 2, 6-dichlorophenolindophenol, ferricyanide, and coenzyme Ql, but not menadione.
D-Gluconate oxidase activity was restored by addition of coenzyme Q2 to a mixture of the purified enzyme and the enzyme-depleted membrane. Coenzyme Q1 or Q4 was also effective to a lesser degree. Coenzyme Q2 appeared to mediate between the enzyme and the membrane residue in an electron transport pathway in which 2, 6-dichlorophenolindophenol or tetramethylphenylenediamine was not involved. The oxidase activity restored by adding coenzyme Q2 was inhibited by cyanide, azide, antimycin A, and n-heptyl-4-hydroxyquinoline-N-oxide. The reconstituted oxidase system possessed a higher optimum pH than the native oxidase system.
Based on these results, the function of each of the three polypeptide components of purified gluconate dehydrogenase is discussed in relation to electron transport in gluconate oxidation.View full abstractDownload PDF (575K) -
Hiroaki SUGINO, Noriyoshi SAKABE, Kiwako SAKABE, Sadashi HATANO, Fumio ...1979 Volume 86 Issue 1 Pages 257-260
Published: July 01, 1979
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSSmooth muscle G-actin from chicken gizzard and Physarum plasmodium G-actin both interact with DNase I and form 1:1 complexes. These complexes were crystallized by using polyethylene glycol 6000 as a precipitant. Both crystals belong to the same orthorhombic space group P212121. The cell dimensions of chicken gizzard G-actin•DNase I complex are a=42.00±0.07 Å, b=225.3±0.4 Å, and c=77.4±0.1 Å, while those of Physarum G-actin•DNase I complex are a=42 Å, b=221 Å, and c=77 Å.View full abstractDownload PDF (736K) -
Flavio MEGGIO, Arianna DONELLA-DEANA, Lorenzo A. PINNA1979 Volume 86 Issue 1 Pages 261-264
Published: July 01, 1979
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSPhosphorylation by a cAMP-independent rat liver protein kinase of protein substrates containing the structural feature required by mammary gland casein kinase (-Ser-X-Glu/Asp) has been demonstrated. In particular, the Bowman-Birk Soybean trypsin inhibitor, which is characterized, like other legume protease inhibitors, by clusters of acidic residues near the C-terminal side of seryl residue(s), proved to be a good model substrate for the protein kinase. Its phosphorylation, involving the Ser 65 residue, is apparently hindered by the binding of trypsin, while it is stimulated by unfolding induced by reduction and subsequent carboxy-methylation.View full abstractDownload PDF (320K) -
Cerebroside Sulfotransferase Activity in Human Lung Tissues An Elevated Level in Lung AdenocarcinomaShinsei GASA, Akira MAKITA, Motohiro HIRAMA, Makoto KAWABATA1979 Volume 86 Issue 1 Pages 265-267
Published: July 01, 1979
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSCerebroside sulfotransferase activity was demonstrated in particulate fractions from human lung and its carcinoma tissues. The activity in human lung adenocarcinoma was significantly higher than those in a different histological type of carcinoma (squamous cell carcinoma) and in the normal tissue from which each carcinoma was derived.View full abstractDownload PDF (222K) -
Sachiko BANDO, Yoshiki MATSUURA, Nobuo TANAKA, Noritake YASUOKA, Masao ...1979 Volume 86 Issue 1 Pages 269-272
Published: July 01, 1979
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSTwo crystalline forms of cytochrome c3 isolated from two strains of Desulfovibrio vulgaris, Miyazaki, tentatively designated as D. vulgaris, Miyazaki F and D. vulgaris, Miyazaki K, have been found. Both belong to the orthorhombic system, space group P212121, but have different cell dimensions; a=54.1, b=68.9, and c=35.0 Å for D. vulgaris, Miyazaki F, and a=43.5, b=41.2, and c=62.9 Å for D. vulgaris, Miyazaki K. The asymmetric unit of each crystal contains one molecule of cytochrome c3.View full abstractDownload PDF (704K) -
Yasuhide SEKI, Makoto ISHIMOTO1979 Volume 86 Issue 1 Pages 273-276
Published: July 01, 1979
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSSirohydrochlorin chromophore prepared by acetone/HCl treatment of desulfoviridin, the sulfite reductase from Desulfovibrio, catalyzed the reduction of sulfite to sulfide and thiosulfate in equimolar amounts when coupled with a hydrogen-hydrogenase-methyl viologen system. This activity was manifested at acidic pH and increased exponentially with decrease in pH. The Km value for sulfite was nearly 10 times that of desulfoviridin. Inorganic iron was necessary for the reduction, since inactivation occurred on passage through a Sephadex LH-20 column or in the presence of 2, 2'-bipyridine, and reactivation was observed on adding iron. The chromophore catalyzed the reduction of dithionite and hydroxylamine.View full abstractDownload PDF (278K) -
Nikolaus SEILER, Thomas SCHMIDT-GLENEWINKEL, Shakir SARHAN1979 Volume 86 Issue 1 Pages 277-278
Published: July 01, 1979
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSγ-Aminobutyric acid is not formed in significant amounts from putrescine by incubation with rat brain homogenates. However, it is formed if acetyl-CoA is added to the incubation medium. This is taken as further evidence for the existence of a metabolic pathway in mammalian brain which comprises acetylation of putrescine to monoacetyl putrescine and oxidative deamination of monoacetyl putrescine by MAO. Nerve cells and glia cells have comparable capacities for putrescine degradation along this pathway.View full abstractDownload PDF (152K) -
1979 Volume 86 Issue 1 Pages 279
Published: 1979
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSDownload PDF (16K)
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