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Volume 77 (1975)
- Issue 6 Pages 1127-
- Issue 5 Pages 909-
- Issue 4 Pages 689-
- Issue 3 Pages 473-
- Issue 2 Pages 265-
- Issue 1 Pages 1-
Volume 77, Issue 3
Displaying 1-24 of 24 articles from this issue
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Takachika AZUMA, Takashi ISOBE, Kozo HAMAGUCHI1975 Volume 77 Issue 3 Pages 473-479
Published: March 25, 1975
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSThe recombination of alkylated H and L chains of a human myeloma protein (Jo) was studied by means of circular dichroism (CD). Marked CD changes were observed at 295 and 235nm when H and L chains recombined. The change in the CD max-imum at 235nm was followed with time after mixing preparations of H and L chains in the pH range between 4 and 6. The recombination reaction was slow and followed second order kinetics. The observed rate constants were markedly dependent on pH. The pH dependence of the rate constant was analyzed assuming that there are two forms of H chain which are in a pH-dependent equilibrium with each other.View full abstractDownload PDF (536K) -
Fumio KISHIDA, Takachika AZUMA, Kozo HAMAGUCHI1975 Volume 77 Issue 3 Pages 481-491
Published: March 25, 1975
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESS1. The proteins precipitated with ammonium sulfate from the urine of a patient (Mat) with multiple myeloma were separated into three components by ion-exchange and gel chromatographies. Sodium dodecyl sulfate polyacrylamide gel electrophoresis, amino acid analyses, immunochemical tests, and measurement of circular dichroism showed that these components were a dimer with a disulfide bond, a stable monomer, and a variable fragment, respectively.
2. All three protein components reacted with 5, 5'-dithiobis-(2-nitrobenzoic acid) in Tris-HCl buffer at pH 8.0, indicating that they contained free sulfhydryl groups. Partial reduction with dithiothreitol in the absence of denaturants yielded two SH groups per molecule from both the monomer and the dimer, and one SH group per molecule from the fragment. This indicates that the monomer of Mat protein contains a cysteinyl residue in the variable region in addition to a cysteinyl residue at the COOH terminus.
3. The reactivities of the two SH groups of the partially reduced monomer toward iodoacetamide and iodoacetic acid were studied by polyacrylamide gel electrophoresis. The two SH groups had similar reactivities with iodoacetamide, but the SH group at the COOH terminus was more reactive with iodoacetic acid than that in the variable region.
4. The extrinsic Cotton effects of an azobenzene-2-sulfenyl group introduced into the SH group in the variable region were different from those of dye attached to the COOH terminal SH group, indicating that the two SH groups had different environments. The states of the SH groups of the intact monomer are discussed on the basis of these findings.View full abstractDownload PDF (3494K) -
Tateo YAMANAKA1975 Volume 77 Issue 3 Pages 493-499
Published: March 25, 1975
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSThe reactivities of cytochromes c with certain redox enzymes were studied. Cow cytochrome oxidase [EC 1. 9. 3. 1], yeast cytochrome c peroxidase [EC 1. 11. 1. 5], and Thiobacillus novellus sulphite-cytochrome c reductase [EC 1. 8. 2. 1] are quite similar to one another in their specificity for cytochrome c. However, all cytochromes c do not react similarly with these enzymes.
Although cytochrome c-555 (Chlorobium thiosulfatophilum) reacts fairly rapidly with the cow oxidase, it does not react either with the yeast peroxidase or the sulphite-cytochrome c reductase. Cytochrome c-550 (Micrococcus denitrificans), cyto-chromes c2 of nonsulphur purple bacteria, and cytochrome c-550 (T. novellus) react much faster with the yeast peroxidase than would be expected from their reactivity with the cow oxidase.
It seems very interesting that the specificity of a reductase (sulphite-cytochrome c reductase) for cytochrome c is similar to that of an oxidase (cytochrome oxidase), since it has been claimed that the reaction sites of the cytochrome c molecule differ between the reductase and oxidase reactions.
The modified cytochrome c-555 (C. thiosulfatophilum), in which three lysine residues (on average) of the molecule are acetylated, reacts with Pseudomonas aeruginosa nitrite reductase [EC 1. 9. 3. 2] as rapidly as the intract cytochrome, while it has lost its reactivity with the cow oxidase completely.View full abstractDownload PDF (557K) -
Partial Purification, Properties, and Subcellular Distribution of Rat Live Phosphatidate PhosphataseKohei HOSAKA, Satoshi YAMASHITA, Shosaku NUMA1975 Volume 77 Issue 3 Pages 501-509
Published: March 25, 1975
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSPhosphatidate phosphatase [EC 3. 1. 3. 4] was purified 15- to 20-fold from the soluble fraction of rat liver. The purification procedure involved calcium phosphate gel adsorption and elution, ammonium sulfate precipitation, and molecular-sieve chroma-tography. For the enzyme assay, an aqueous dispersion of phosphatidate, rather than “membrane-bound” phosphatidate, was used as substrate.
The partially purified enzyme depends almost entirely on the presence of Mg2+ for its activity. Moreover, the activity of the enzyme is stimulated by phosphatidyl-choline. The enzyme exhibits a high substrate specificity for phosphatidate. The apparent Km for phosphatidate is approximately 0.05mM. The optimum pH is between 7.4 and 7.6. The enzyme is inhibited by fluoride and by p-chloromercuri-benzoate.
The subcellular distribution of phosphatidate phosphatase in rat liver was studied by assaying the activity of the enzyme in the presence of Mg2+ and phosphatidyl-choline. In contrast to the results of previous studies, most of the enzyme activity was found in the soluble fraction.View full abstractDownload PDF (745K) -
Kaoru TSUJII, Toshio TAKAGI1975 Volume 77 Issue 3 Pages 511-519
Published: March 25, 1975
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSThe binding of an anionic surfactant to a protein polypeptide has been studied by the proton magnetic resonance (PMR) technique to form a part of our studies on the principles of SDS-polyacrylamide gel electrophoresis. Sodium 4-(p-butylphenyl) butane-1-sulfonate (CH3-(CH2)3-φ-(CH2)4-SO3-Na+) was employed as an anionic surfactant, and reduced and carboxyamidomethylated (RCAM) bovine serum albumin as a typical protein polypeptide. The binding isotherm of the surfactant to RCAM bovine serum albumin was similar to that of sodium dodecyl sulfate (SDS). The surfactant could replace SDS in SDS-polyacrylamide gel electrophoresis without affecting the well-known mode of separation of protein bands. These results gave a sound basis for the assumption that the investigation of the complex between a surfactant with a benzene ring and RCAM bovine serum albumin would provide useful knowledge concerning the principles of SDS-polyacrylamide gel electrophoresis.
Aggregation of the aromatic surfactant necessarily brings benzene rings together. A benzene ring is a strong source of the ring current effect on chemical shifts in nuclear magnetic resonance (NMR). Chemical shifts of the surfactant in NMR are, therefore, sensitive to whether the surfactant molecules are single-molecularly dis-solved or aggregated. Full advantage was taken of the above fact in the present PMR study of the binding of the surfactant to RCAM bovine serum albumin.
The chemical shifts of the phenyl and methyl protons both for the single-molecular and micellar aggregated states were estimated from measurements of the shifts as a function of the surfactant concentration. They shifted to a higher magnetic field on micelle formation, due to the increase of the ring current effect.View full abstractDownload PDF (1600K) -
Yoshihiro FUKUSHIMA, Yuji TONOMURA1975 Volume 77 Issue 3 Pages 521-531
Published: March 25, 1975
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSThe effects of Na+ and K+ ions on the elementary steps in the reaction of Na+-K+-dependent ATPase [EC 3. 6. 1. 3] were investigated in 0.5-600mm NaCl and 0-10mM KCl, at a fixed concentration (1mM) of MgCl2, at pH 8.5 and at 15°. The data were analyzed on the basis of the reaction mechanism in which a phosphorylated inter-mediate, _??_(abbreviated as EP), is formed via two kinds of enzyme-substrate complex, E1ATP and E2ATP, and EP is in equilibrium with E2ATP, and is hydrolyzed to produce P1 and ADP.
The following results were obtained:
1. The rate of E2ATP-formation, vf, increased with increase in the Na+ concentration, reached a maximum level, and then decreased with further increase in the Na+ concentration at various K+ concentrations. The value of vf was given as_??_, where_??_, and_??_
2. The reciprocal of the equilibrium constants, K3, of the step E1ATP_??_in the presence of low concentrations of Na+ was larger than that in the presence of high concentrations of Na+, indicating that the equilibrium shifted markedly toward E2ATP at low concentrations of Na+. The relation of K3 with Na concentra-View full abstractDownload PDF (708K) -
Yoshihiro FUKUSHIMA, Yuji TONOMURA1975 Volume 77 Issue 3 Pages 533-541
Published: March 25, 1975
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSPreviously, we proposed the following reaction machanism for the transport ATPase [EC 3. 6. 1. 3] reaction in the presence of high concentrations of Mg2+ and Na+:
E+ATP_??_E1ATP→E2ATP_??_→E+ADP+P1. step 1 2 3 4
Some kinetic and thermodynamic properties of steps 3 and 4 were investigated, and the following results were obtained.
1. When the reaction was started by adding ATP to the enzyme in the presence of 50mM Na+ and 0.5mM K+ or in the presence of 50mM Na+ and 0.5mM Rb+, the amount of_??_increased with time and maintained a constant level after reaching a maximum. We could not observe the initial burst of EP formation, which was observed by Post et al. in the presence of 8mM Na+ and 0.01mM Rb+.
2. The existence of quasi-equilibrium between E2ATP and_??_in the presence of low concentrations of Na+ was suggested by the fact that the values of the reciprocal of the equilibrium constant, K3, of step 3 obtained by the following three methods were almost the same. a) The value of 1+K3 was estimated from the ratio of v0/[EP] to kd, where v0 is the rate of ATP hydrolysis in the steady state, [EP] the concentration of EP, and kd the first-order rate constant of EP disappearance after stopping EP formation. b) This value was also calculated from the ratio of the amount of Pi liberated to that of decrease in EP after stopping EP formation. c) The value of K3 was also calculated from the initial rapid decrease in EP on adding K+ and EDTA, assuming that the rapid decrease was due to a shift of the equilibrium toward E2ATP on adding K+. For example, the value of K3 with 10mM NaCl andView full abstractDownload PDF (624K) -
Relation between the Extent of Desensitization and the Amount of Bound p-ChloromercuribenzoateKazuko SHIBATA-SEKIYA, Yuji TONOMURA1975 Volume 77 Issue 3 Pages 543-557
Published: March 25, 1975
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSH-Meromyosin (CMB→βME-H-meromyosin) was prepared by tryptic digestion of myosin, which had been treated with CMB and then β-mercaptoethanol. The relation between the amount of CMB bound to H-meromyosin and the extent of desensitization of the substrate inhibition of acto-H-meromyosin ATPase [EC 3. 6. 1. 3] was investigated.
Both the dissociation of acto-H-meromyosin induced by ATP and substrate inhibition decreased with increase in the amount of bound CMB to a minimum value at about 1 mole of CMB bound per mole of H-meromyosin. The substrate inhibition of acto-H-meromyosin ATPase was restored to the original level by complete removal of the bound CMB by further treatment of CMB→βME-H-meromyosin with a large excess of β-mercaptoethanol.
The dissociation constant of acto-H-meromyosin in the presence of ATP decreased markedly on modification with CMB, while the maximum ATPase activity at a sufficiently high concentration of F-actin remained essentially unchanged. Acto-H-meromyosin was reconstituted from F-actin and CMB→βME-H-meromyosin, containing less than the stoichiometric amount of bound CMB. Its ATPase activity and the extent of dissociation of acto-H-meromyosin induced by ATP were explained as those of a mixture of unmodified H-meromyosin and CMB→βME-H-meromyosin containing 1 mole of CMB per mole of H-meromyosin.
Half of the light chains (g2), with a molecular weight of 18, 000, were removed from myosin by treatment with CMB and β-mercaptoethanol. After this treatment, on further incubation of the myosin with a large excess of β-mercaptoethanol, the myosin contained only half of the g2, but the substrate inhibition of acto-H-meromyosin ATPase was restored completely.View full abstractDownload PDF (1137K) -
Koichiro TAKESHIGE, Akira NAKAGAWARA, Tanenori HATAE, Shigeki MINAKAMI1975 Volume 77 Issue 3 Pages 559-566
Published: March 25, 1975
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSThe addition of trypsin [EC 3. 4. 21. 4]-digested liver microsomes induced cyanide-insensitive respiration in guinea pig polymorphonuclear leucocytes with concomitant acceleration of the hexose monophosphate oxidative pathway. The respiration was insensitive to inhibitors of mitochondrial respiration but sensitive to glycolytic inhibitors. These metabolic alterations are similar to those associated with phagocytosis, though the digested microsomes were apparently not taken up by the cells and probably trigger the metabolic changes by interaction with the cellular membrane. Intact microsomes or microsomes treated with chymotrypsin [EC 3. 4. 21. 1], bacterial proteinase, ribonuclease [EC 3. 1. 4. 22], or neuraminidase [EC 3. 2. 1. 18] could not induce such respiration.View full abstractDownload PDF (1456K) -
Akira NAKAGAWARA, Koichiro TAKESHIGE, Shigeki MINAKAMI1975 Volume 77 Issue 3 Pages 567-573
Published: March 25, 1975
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSThe effects of N6, O2'-dibutyryladenosine 3', 5'-monophosphate (Bu2-cyclic AMP) and N2, O2'-dibutyrylguanosine 3', 5'-monophosphate (Bu2-cyclic GMP) on the cyanide-insensitive respiration of guinea pig peritoneal exudate polymorphonuclear leucocytes were studied. Bu2-cyclic AMP inhibited the respiration induced both by phagocytosis of E. coli and by the interaction with trypsin-digested rat liver microsomes. The addition of theophylline gave rise to an inhibitory pattern similar to that with Bu2-cyclic AMP against both the respirations induced. On the other hand, Bu2-cyclic GMP did not affect the respiration induced by phagocytosis whereas it inhibited the respiration induced by trypsin-digested microsomes. The cyanide-insensitive respiration induced by the addition of myristic acid was inhibited by Bu2-cyclic AMP, which was similar to that with E. coli. The respiration induced by methylene blue was inhibited neither by Bu2-cyclic AMP nor by Bu2-cyclic GMP. These observations suggest that the cyanide-insensitive respiration of polymorphonuclear leucocytes may be classified into at least three types from the inhibitory pattern of cyclic AMP and cyclic GMP.View full abstractDownload PDF (460K) -
Akinori AMEMURA, Hironoshin KITAGAWA, Tokuya HARADA1975 Volume 77 Issue 3 Pages 575-578
Published: March 25, 1975
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESS1. Pullulanase [EC 3. 2. 1. 41] was inhibited by Hg2+, N-bromosuccinimide, 2-hydroxy-5-nitrobenzyl bromide, riboflavine 5'-phosphate, histamine, imidazole, and carbodiimide.
2. The tryptophan groups of the pullulanase were modified by N-bromosuccinimide. It was found that one or two of the 25 tryptophan groups seemed to be important for the activity.
3. Studies on difference spectra using various substrates suggested that the tryptophan groups were important for the formation of an enzyme-substrate complex.View full abstractDownload PDF (267K) -
Khaik Cheang OO1975 Volume 77 Issue 3 Pages 579-586
Published: March 25, 1975
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSA particulate enzyme preparation from Bacillus stearothermophilus synthesized 1, 3-poly (glycerol phosphate) from CDPglycerol at an optimum pH of 8.0 and the reaction was stimulated by divalent cations. Km for CDPglycerol was 0.18mM. The synthesis was inhibited by CMP, CDP, and CTP and by concentrations of CDP-glycerol above 0.49mM. The reaction was irreversible. The product had an average chain length of 8 glycerol units. About two thirds of the polymers were synthesized in entirety while the remainder were attached to some acceptor by their phosphate end. The enzyme was able to synthesize only a limited amount of polymer.View full abstractDownload PDF (644K) -
Koichi SUZUKI, J. Ieuan HARRIS1975 Volume 77 Issue 3 Pages 587-593
Published: March 25, 1975
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESS1. Glyceraldehyde-3-phosphate dehydrogenases [EC 1. 2. 1. 12] from rabbit, pig, lobster, yeast, E. coli, and B. stearothermophilus have been subjected to hybridization in 3M NaCl.
2. Suitable mixtures of electrophoretically distinct glyceraldehyde-3-phosphate dehydrogenases were found to give five-membered hybrid sets, revealed by protein as well as by activity staining, when examined by electrophoresis on cellulose acetate.
3. The thermophile enzyme did not form hybrids with any of its mesophile counter-parts, presumably because it does not dissociate under the conditions used.
4. Hybridization of pig enzyme with lobster enzyme that had been inactivated by selective carboxymethylation of Cys-149 gave rise to four enzymatically active protein bands, including the tetramer containing only one active pig enzyme subunit. Individual subunits would thus appear to express their activity independently even within hybrid tetramers formed with subunits of another species.
5. The successful hybridization of glyceraldehyde-3-phosphate dehydrogenases from evolutionarily distant sources suggests that the tertiary and quaternary structures of the enzyme have been highly conserved.View full abstractDownload PDF (2331K) -
Shiroki YAGISAWA1975 Volume 77 Issue 3 Pages 595-604
Published: March 25, 1975
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSOne mole of horse hemoglobin tetramer reacts with 2 moles of 2-chloromercuri-4-nitrophenol (MNP) at β93 cysteine. The difference spectra between MNP-bound hemoglobin and hemoglobin, measured with the aid of ascorbic acid and ascorate oxidase [EC 1. 10. 3. 3] as deoxygenation reagents, indicate that the pK of the phenolic hydroxyl group of MNP increases by 0.6 to 0.8 pH unit on deoxygenation of the hemoglobin. The Hill constant of the modified hemoglobin changes with pH. It decreases from about 2.4 at pH 6.8 to about 1.0 at pH 9.0. This effect of the reagent is interpreted as inherent to the reporter groups.View full abstractDownload PDF (793K) -
Shiroki YAGISAWA1975 Volume 77 Issue 3 Pages 605-616
Published: March 25, 1975
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSA method is proposed for observation of the interaction between charged macro-molecules such as proteins. The method is based on the fact that the pK of an ionizable reporter group attached to a macromolecule can be altered by the electro-static effect of another charged macromolecule which associates with the former. The effectiveness of the method was shown in the study of the association of bovine serum albumin with hen egg lysozyme [EC 3. 2. 1. 17]. The errors inherent in this method in obtaining the equilibrium constant of the association reaction and procedures for their correction are discussed.View full abstractDownload PDF (853K) -
Masaaki MASUBUCHI, Shinji MIURA, Zensaku YOSIZAWA1975 Volume 77 Issue 3 Pages 617-626
Published: March 25, 1975
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESS1. Complex carbohydrate fractions were extracted successively with 40% aqueous EDTA (pH 7.4) and 6M urea (pH 7.8) from acetone-dried bone powder of rabbit femur.
2. The carbohydrate fraction extracted with EDTA (E-Fr) was separated into five fractions, Dl_??_D5, by DEAE-Sephadex A-50 column chromatography. Chemical and infrared spectral analyses, and enzymatic digestion indicated that D2 contained less-acidic glycoprotein, D3 contained sialoglycoprotein, D4 contained a low sulfated proteokeratan sulfate-like substance, and D5 contained glycoprotein-bound chondroitin sulfate A plus protein-free chondroitin sulfate A.
3. Two fractions, HU-D1 and HU-D2, were isolated from the carbohydrate fraction extracted with urea (HU-Fr) by successive digestion with collagenase [EC 3. 4. 99. 5] and pronase, followed by gel-filtration on Sephadex G-100 and then DEAE-Sephadex A-50 column chromatography. HU-D1 and HU-D2 contained a low sulfated keratan sulfate-like substance linked to peptide and glycopeptide-bound chondroitin sulfate A, respectively.
4. The present findings indicate that rabbit femur contains low sulfated proteo-keratan sulfate-like substances with varying sulfate contents and glycoprotein-bound chondroitin sulfate A as the principal glycosaminoglycans. The macromolecules bound more tightly to the tissue contain much more sulfate than the corresponding loosely bound ones.View full abstractDownload PDF (720K) -
Teruo MATSUBARA1975 Volume 77 Issue 3 Pages 627-632
Published: March 25, 1975
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSThe oxidation of cytochromes during the reduction of N2O to N2 by a denitrifying bacterium was studied spectrophotometrically.
The reduced b- and c-type cytochromes are partially oxidized when N2O is added to intact cells reduced with lactate under anaerobic conditions. The oxidation of cytochromes is inhibited non-competitively by azide, cyanide, 2, 4-dinitrophenol and CuSO4, which inhibit the reduction of N2O to N2. In the presence of each inhibitor at a high concentration, at which the reduction of N2O to N2 is perfectly inhibited, cytochromes are not oxidized by N2O, while when an adequate, low concentration of inhibitor is added, b-type cytochrome is partially oxidized but c-type cytochrome is apparently not oxidized. In cell-free extracts, prepared by the sonic disruption of cells, that have entirely lost their activity in the reduction of N2O to N2, cytochromes are not oxidized by N2O.
From the above results, it was concluded that b-type and c-type cytochromes should participate in the electron transport mechanism of the reduction of N2O to N2.View full abstractDownload PDF (478K) -
Iwao OHTSUKI1975 Volume 77 Issue 3 Pages 633-639
Published: March 25, 1975
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESS1. The localization of specific antibodies against troponin components, i.e., troponin T (TN-T), troponin I (TN-I), and troponin C (TN-C), was studied by the use of an electron microscope.
2. Every antibody was distributed along the thin filament with a period of 38nm.
3. Staining with anti-TN-I or anti-TN-C formed narrow striations. The location of the first striation was 26nm from the free end of the thin filament.
4. The width of individual striations formed by anti-TN-T was 14-20nm. The H-band-side end of each striation coincided with the location of anti-TN-I or anti-TN-C.View full abstractDownload PDF (4090K) -
Yosuke EBINA, Hajime IWAI, Noriko FUKUI, Hidesato OHTSUKA, Yoshiaki MI ...1975 Volume 77 Issue 3 Pages 641-645
Published: March 25, 1975
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSAfter partial hepatectomy in rats, the following changes in enzymic activities were observed in the remnant liver during the prereplicative period. In the initial period of the prereplicative process, soon after removal of part of the liver, ornithine decarboxylase [EC 4. 1. 1. 17] and IMP dehydrogenase [EC 1. 2. 1. 14] increase. Sub-sequently, for entry into the S period, thymidine kinase [EC 2. 7. 1. 75] increases simultaneously with increase in the intracellular cyclic AMP level and decrease in its phosphodiesterase [EC 3. 1. 4. 17].View full abstractDownload PDF (375K) -
Takao ARAI, Yasuo IHARA, Ken-ichi ARAI, Yoshito KAZIRO1975 Volume 77 Issue 3 Pages 647-658
Published: March 25, 1975
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSA rapid and sensitive assay for [3H] GTP binding activity of tubulin has been developed. This assay method is based on the quantitative retention of [3H] GTP. tubulin complex on a nitrocellulose membrane filter. It was also found that bovine brain tubulin is markedly stabilized by glycerol and GTP against denaturation. A large-scale purification of bovine brain tubulin was achieved using the new assay procedure and by the inclusion of glycerol and GTP in a buffer solution used for column chromatography. The purified tubulin could be stored at -80° in the presence of glycerol and GTP for at least a year without any appreciable loss of [3H] GTP- and [3H] colchicine binding activities.
The interaction of tubulin with guanine nucleotides was also studied using the nitrocellulose membrane filter procedure. It was found that the binding of [3H] GTP to tubulin with an empty exchangeable site proceeded promptly within 5 sec while the exchange of [3H] GTP with a GTP•tubulin complex in which the exchangeable site had been occupied with unlabeled GTP occurred more slowly. The dissociation constants for GTP and GDP at the exchangeable site of tubulin were determined as 0.5×10-6M and 1.9×10-6M, respectively. 5'-Guanylylimidodiphosphate could interact, although less strongly, with tubulin at this site, whereas the interaction of other nucleoside triphosphates including ATP, CTP, UTP, and 5'-guanylyl methylenedi-phosphonate was very weak, if it occured at all. The presence of Mg2+ and a free sulfhydryl group was found to be essential for binding of [3H] GTP to tubulin. Ca2+ was found to replace Mg2+ in this binding reaction.View full abstractDownload PDF (1005K) -
I. Urea Synthesis with Ammonia and Glutamine as Nitrogen SourcesTakeyori SAHEKI, Nobuhiko KATUNUMA1975 Volume 77 Issue 3 Pages 659-669
Published: March 25, 1975
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSUrea synthesis was studied using the isolated liver perfusion system with ammonium chloride and glutamine as nitrogen sources. The rate of urea formation increases with ammonium chloride concentration up to 5mM, and the rate remained constant in the range between 5 and 20mM of ammonium chloride as the substrate. The concentration of ammonia in the medium to support the half-maximum velocity of urea formation was 0.7mM. The rate of urea formation was stimulated by the addition of 2.5mM ornithine, and the greater part of the ornithine which was taken up into the liver was accumulated as citrulline in the presence of ammonia. A considerable accelerating effect of N-acetylglutamate on the synthetic rate was observed, but a rather high concentration of N-acetylglutamate was required in order to obtain the maximum effect possibly, because its permeability into liver cells may be limited. A marked additive effect on the rate of urea formation was observed with the combined addition of ornithine and N-acetylglutamate.
The metabolic conversion of glutamine nitrogen to urea in the perfused rat liver and the effect of several compounds which stimulated urea synthesis with ammonia were further examined. The process of conversion of glutamine nitrogen to urea might be composed of the following three steps. In the first lag phase, a small amount of glutamine was removed from the medium. In the second stage, the glutamine level decreased rapidly and ammonia was accumulated in the perfusate. The third stage was a period in which glutamine concentration remained at a con-stant low level, and the accumulated ammonia was rapidly conversed to urea. The rate of urea formation in this third stage was found to be much higher than that with ammonia as the substrate. The maximum rate of glutamine removal was obtained at pH 7.7 of the perfusate and at a concentration of 10mM glutamine.
Urea formation with glutamine was also stimulated by the addition of ornithine, malate, or N-acetylglutamate, which had accelerating effects on the urea synthesis with ammonia. This stimulation was due to an effective conversion of ammonia to urea, but no change in the rate of removal of glutamine was obtained.View full abstractDownload PDF (895K) -
II. Comparison of Urea Synthesis in Livers of Rats Subjected to Different Dietary ConditionsTakeyori SAHEKI, Michio TSUDA, Tomi TANAKA, Nobuhiko KATUNUMA1975 Volume 77 Issue 3 Pages 671-678
Published: March 25, 1975
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSCapacities for urea synthesis and amino acid patterns in the perfused livers isolated from rats fed low and high-protein diets were compared.
Urea formation with ammonium chloride as the nitrogen source in perfused livers isolated from rats fed on a 70% casein diet was rapid and the efficiency of conversion of ammonia to urea was 97.9%. However, that in livers isolated from rats fed on a 5% casein diet was much slower and the efficiency of conversion of ammonia to urea was only 36.1%.
The ratios of the rate of urea formation from ammonium chloride to activity of ornithine transcarbamylase [EC 2. 1. 3. 3] in the perfused livers of rats fed on 5 and 70% casein diets were calculated. The ratio of the former condition was much lower than that of the latter. The ratios reached nearly the same level by the addition of ornithine and N-acetylglutamate, the addition of which to the perfusate caused marked elevation of the ratios in both cases.
In the perfused livers from rats fed on a 5% casein diet a considerable portion of the ammonia added to the perfusate was fixed into an amino or an amide group of amino acids such as alanine, aspartate, and glutamine. On the other hand, in the perfused livers from rats fed on a 70% casein diet most of the ammonia added was converted to urea. The regulation of urea synthesis and the relation between anabolism and catabolism of amino acids in rat livers subjected to different dietary conditions were compared.View full abstractDownload PDF (594K) -
Yasuhiro TAKEDA, Makoto KAGEYAMA1975 Volume 77 Issue 3 Pages 679-684
Published: March 25, 1975
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSAn optical diffraction study has been made of electron micrographs of extended sheaths of pyocin R. It has been demonstrated that the extended sheath consists of annuli of six subunits, which are arranged in a helix of 3.57 annuli per one turn. The annulus repeat in the helix direction is 35 A. The dimensions and probably the helical parameters of pyocin R sheath are different from those of T4 phage tail, but a structural correlation appears to exist between the extended sheath of pyocin R and that of the phage tail.View full abstractDownload PDF (3255K) -
Yahito KOTAKE, Takashi UEDA, Terutoshi MORI, Edahiko MURAKAMI, Masayas ...1975 Volume 77 Issue 3 Pages 685-687
Published: March 25, 1975
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSThe xanthurenic acid-insulin complex was found to have similar immunological properties to native Zn-insulin. This complex showed less hormonal activity on glucose metabolism in adipose tissue than native Zn-insulin, but its activity was increased by addition of Zn2+ ions.View full abstractDownload PDF (215K)
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