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Volume 92 (1982)
- Issue 6 Pages 1683-
- Issue 5 Pages 1337-
- Issue 4 Pages 977-
- Issue 3 Pages 607-
- Issue 2 Pages 319-
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Volume 92, Issue 4
Displaying 1-41 of 41 articles from this issue
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Fumi MORITA, Shuhei KONDO1982 Volume 92 Issue 4 Pages 977-983
Published: October 01, 1982
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSMyosin purified from the smooth muscle of scallop adductor contains two kinds of regulatory light chain, regulatory light chain a (RLC-a) and regulatory light chain b (RLC-b) (Kondo, S. & Morita, F. (1981) J. Biochem. 90, 673). The myosin was fractionated by salting out with ammonium sulfate, and samples containing the two regulatory light chains with different molar ratios were obtained. ATPase activities of the myosin fractions were determined. From the analysis of the de-pendence of ATPase activity on molar ratio of the two regulatory light chains, we concluded that myosin purified from the smooth muscle of scallop contains three species of myosin having different combinations of regulatory light chains: one has two RLC-a (aa), another has two RLC-b (bb), and the third one each of RLC-a and RLC-b (ab). The order of ATPase activities of these three myosin species was estimated as (aa)<(bb)<(ab).
Distribution of the two regulatory light chains in the smooth muscle from the inside, translucent portion to the outside, opaque portion was examined by means of one- and two-dimensional gel electrophoreses. The content of RLC-b was about 1 mol per mol of SH-light chain independent of the portion of muscle. The content of RLC-a was markedly dependent on the portion of muscle-about 0.2 mol per mol of SH-light chain in the innermost portion and 0.7 mol per mol of SH-light chain in the outside, opaque portion. The sum of both regulatory light chain contents was about 1.5 mol per mol of SH-light chain in the opaque portion where the catch contraction is notable. Myosin species in the catch muscle are discussed.View full abstractDownload PDF (1107K) -
Hajime TAKIKAWA, Hideaki OTSUKA, Tomoe BEPPU, Yousuke SEYAMA1982 Volume 92 Issue 4 Pages 985-998
Published: October 01, 1982
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSIndividual non-glucuronidated•non-sulfated, glucuronidated and sulfated bile acids in serum were determined, i.e. lithocholic acid, deoxycholic acid, chenodeoxycholic acid, ursodeoxycholic acid, and cholic acid, by mass fragmentography. Glucuronic acid conjugates of lithocholic acid, deoxycholic acid, chenodeoxycholic acid, and cholic acid were synthesized via the Koenigs-Knorr condensation reaction. Deute-rium labeled deoxycholic acid, lithocholic acid glucuronide, deoxycholic acid glucu-ronide, and deoxycholic acid sulfate were synthesized and used as internal standards. A serum sample of 1 ml including internal standards was purified with a Sep-Pak C18 cartridge. After the enzymatic cleavage of amino acid conjugates, bile acids were separated into three fractions, free, glucuronidated, and sulfated bile acids, using piperidinohydroxypropyl Sephadex LH-20 (Goto et al. (1978) Clin. Chim. Acta 87). Glucuronidated and sulfated bile acids were deconjugated by β-glucuronidase treat-ment and solvolysis. Each fraction was converted to the hexafluoroisopropyl-tri-fluoroacetyl derivative and quantitated by mass fragmentography. The average concentrations of individual bile acid glucuronides from healthy fasting subjects (n=9) were as follows; lithocholic acid 0.013μg/ml, deoxycholic acid 0.083μg/ml, chenodeoxycholic acid 0.078μg/ml, ursodeoxycholic acid 0.013μg/ml, and cholic acid 0.007μg/ml. Bile acid glucuronides occupied 7.8% of the total bile acids.View full abstractDownload PDF (1770K) -
Tohru AKUTAGAWA, Tatsuo OOI1982 Volume 92 Issue 4 Pages 999-1007
Published: October 01, 1982
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSLight meromyosin (LMM) prepared from rabbit skeletal myosin (with trypsin (LMM (T)) or with chymotrypsin (LMM (CT)) was digested with trypsin and chymo-trypsin in order to identify structurally loose regions and regions responsible for the low solubility of LMM at low salt concentrations. Four chains, FT 1 to FT 4, were found in SDS gel electrophoretic patterns of the tryptic digests of LMM (T), their apparent molecular weights being 70, 000, 62, 000, 55, 000, and 47, 000, respec-tively. Among them, FT 1 (corresponding to the undigested LMM chain) and FT 3 were found in a precipitate fragment (FTP) produced at 0.08M KCl pH 7.0, where LMM is insoluble, while FT 2 and FT 4 were the major components in the soluble material (FTS). Since sedimentation experiments showed that LMM (T), FTP, and FTS in 0.6M KCl migrated as single peaks with approximately the same sedimentation velocities (about 3.0 S) and CD measurements revealed that they had α-helical contents of 82, 95, and 94%, respectively, the rodlike molecules in solution are deduced to be composed of two chains, which were observed in SDS gel electro-phoresis. A preparation of LMM with chymotryptic digestion gave a smaller size of LMM (LMM (CT)) (a chain which migrated at the same rate as that of FT 2), which could also be obtained by chymotryptic digestion of LMM (T). Thus, certain limited regions in the subunit chain of LMM are responsible for the low solubility, since one molecule (containing FT 2) is soluble at 0.08M KCl, whereas another molecule of the same size (LMM (CT)) is not. Reconstitution experiments in which a mixture of LMM and soluble fragments were denatured by incubation in 5M guanidine HCl, and the dissociated chains were reassociated after removal of the denaturant, gave further information on the location of the regions. That is, recon-stituted molecules which formed a precipitate at 0.08M KCl consisted of one chain of LMM and one chain of FTS (FT 2 or FT 4), indicating that at least one region in LMM is necessary for the characteristic solubility of LMM. From these results, the primary proteolytic sites are deduced to be located about 8, 000 daltons from the N- and C-termini of LMM, respectively.View full abstractDownload PDF (3270K) -
Nanaya TAMAKI, Hidenori AOYAMA, Keiko KUBO, Tatsuhiko IKEDA, Takao HAM ...1982 Volume 92 Issue 4 Pages 1009-1017
Published: October 01, 1982
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSβ-Alanine aminotransferase from rabbit liver has been purified 1, 700-fold over the initial liver extract. The purified enzyme was shown to be homogeneous by disc electrophoresis and SDS polyacrylamide electrophoresis. The molecular weight of the purified enzyme determined by gel filtration was 95, 000±5, 700 and the subunit molecular weight was 48, 000±2, 100. The enzyme showed absorption maxima at 282, 330, and 414 nm and contained only 1 mol of pyridoxal 5'-phosphate/mol of dimer. The pH optimum for enzyme activity was 8.8 and the Km values for β-alanine and 2-oxoglutaric acid were calculated to be 3.9 and 1.4mM, respectively. The enzyme catalyzed transamination of various ω-amino acids with 2-oxoglutaric acid, which was a favourable amino acceptor. β-Alanine, γ-aminobutyric acid, and β-aminoisobutyric acid, which are naturally occurring substrates, were preferred amino donors, but taurine, alanine, ornithine, spermine, and spermidine were not. 6-Azauracil inhibited the enzyme activity with a Ki of approximately 1.5mM.
From the above properties, β-alanine aminotransferase from rabbit liver was seen to closely resemble with 4-aminobutyrate aminotransferase from liver and brain.View full abstractDownload PDF (1215K) -
Mamoru TAMURA, Nozomu OSHINO, Britton CHANCE1982 Volume 92 Issue 4 Pages 1019-1031
Published: October 01, 1982
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSThe effect of hydroperoxides on the cardiac tissue was studied by using hemoglobinfree perfused rat heart. Ethylhydroperoxide was degraded mainly through the glutathione peroxidase system of the heart at a maximal rate of about 1.2μmol/min per g wet wt. When ethylhydroperoxide infused was not degraded completely, the hydroperoxide concentration in the effluent perfusate paralleled the formation of ferrylmyoglobin in the heart. The infusion of ethylhydroperoxide caused release of oxidized glutathione into the effluent perfusate as a result of the enhancement of the cytosolic glutathione peroxidase reaction. The leakage of oxidized glutathione reached the maximal rate of 3.5 nmol/min per g wet wt with the infusion of 175μM ethylhydroperoxide. At hydroperoxide concentrations above 150 μM, oxidations of pyridine nucleotides and of cytochrome α+α3 occurred, probably through a stimulation of the mitochondrial glutathione peroxidase reaction, and resulted in sudden failure of the heart function. The infusion of t-butyl- and cumene-hydro-peroxides, which are unable to react with myoglobin, also caused the oxidations of pyridine nucleotides and cytochrome α+α3, the inhibition of oxygen consumption and the failure of heart function. The results indicate that the cardiac toxicity of hydroperoxides is due mainly to their effect on mitochondrial metabolism.View full abstractDownload PDF (1070K) -
A Simple Procedure for Purification of NADH-Specific Dihydropteridine Reductase from Mammalian LiverNobuo NAKANISI, Kei HIRAYAMA, Shozo YAMADA1982 Volume 92 Issue 4 Pages 1033-1040
Published: October 01, 1982
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSA simple and convenient purification method which can yield a homogeneous prep-aration from even a small amount of starting material was devised for NADH-specific dihydropteridine reductase from rat liver. The procedure is essentially composed of two steps, i.e., affinity chromatography on Matrex gel blue A and hydrophobic chromatography on Phenyl-Sepharose. Prior to the Matrex gel blue A chromatography, the crude extract of rat liver was oxidized to dissociate NADH, bound in the enzyme-NADH complex, from the enzyme. Low molecular weight substances in the extract were removed by Sephadex G-25 gel filtration; then the enzyme was purified by successive chromatographies on Matrex gel blue A and Phenyl-Sepharose columns. Thus about 0.1mg of purified enzyme was obtained from 3g of rat liver with 40% recovery. The preparation showed a single protein band on polyacrylamide and SDS-gel electrophoresis. Using NADH and tetra-hydro-6-methylpterin, the maximal velocity of the enzyme was determined to be 64.2μmol quinonoid-dihydro-6-methylpterin reduced/min/mg. Km values of the enzyme were 0.85 μM and 3.4 μM for NADH and quinonoid-dihydro-6-methylpterin, respectively. This simple purification method was also applicable to livers from other mammalian sources such as human and bovine.View full abstractDownload PDF (1318K) -
Shiro KAKIUCHI, Seiji YASUDA, Reiko YAMAZAKI, Yoshiko TESHIMA, Keiko K ...1982 Volume 92 Issue 4 Pages 1041-1048
Published: October 01, 1982
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSAlthough calmodulin is generally regarded as a soluble protein, a considerable amount of calmodulin activity was found to be associated with particulate fractions of mammalian tissues after an extensive washing of the particulate fraction with EGTA. Identity of this particle-bound and EGTA-nonextractable form of cal-modulin with soluble calmodulin was established recently (Sobue, K., Yamazaki, R., Yasuda, S., & Kakiuchi, S. (1981) FEES Lett. 129, 215-219). The particle-associated calmodulin activity was latent to some extent and its unmasking required the presence of nonionic detergent. We have developed an assay method for the soluble and particulate forms of calmodulin in biological samples and, by means of this method, concentrations of calmodulin in rat and bovine tissues were quan-titatively determined. In the supernatant, high levels (>10μM) of calmodulin were found in the testis, pituitary gland, and various areas of brain, intermediate levels (5-10μM) in the lung, prostate, and adrenal gland, and low levels (<5μm), in the liver, kidney, and spleen. Particulate fractions contained 10-50% of the total calmodulin contents in the tissues. Human erythrocytes contained (2.5±0.2)μM calmodulin, or (14±0.9)×104 calmodulin molecules per cell.View full abstractDownload PDF (635K) -
Hideyo HASUMI1982 Volume 92 Issue 4 Pages 1049-1057
Published: October 01, 1982
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSThe whole protein structure and the microenvironments of the iron-sulfur cluster and of the side chains of amino acid residues of spinach ferredoxin were studied by optical absorption and circular dichroism (CD) spectroscopy in the alkaline pH range.
From the pH-dependence of the optical absorption changes at 245 nm, the four tyrosyl residues of ferredoxin were classified into three groups: one exposed residue with a normal apparent pK value of 10.1, two exposed residues with abnormal apparent pK values of 12.0, and one buried residue showing time-dependent ionization. The absorption in the visible region disappeared gradually with the ionization of the buried residue rather than that of the three exposed residues. The apparent pK value of 10.0 was obtained from the rapid CD changes at 258 nm caused by pH elevation from neutral to alkaline pH. The structural alteration associated with the CD change had no effect on the secondary structure of the protein moiety other than the iron-sulfur cluster and the microenvironment of the cluster. The rate constants obtained from the time courses of the CD changes in the near-ultraviolet and visible regions were in good agreement with those obtained from the time courses of the optical absorption changes.
These results lead to the conclusions that (1) the native ferredoxin structure is maintained through the interaction with the iron-sulfur cluster and (2) the protein structure in the neighborhood of the cluster, important for the physiological activity, is not perturbed even though the exposed tyrosyl residues are ionized.View full abstractDownload PDF (666K) -
Fumio KISHI, Yousuke EBINA, Toru MIKI, Teruko NAKAZAWA, Atsushi NAKAZA ...1982 Volume 92 Issue 4 Pages 1059-1068
Published: October 01, 1982
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSFrom the cells of an Escherichia coli K-12 strain, a 22, 000-dalton protein which has an affinity for the superhelical DNA molecule was purified to apparent homogeneity by monitoring the DNA-binding activity using the filter binding assay. In the sedimentation analysis of the DNA-protein complex, the protein has an affinity for the superhelical or single-stranded DNA molecule but neither for the open-circular nor for the linear DNA molecule. The amino acid composition of the protein resembled those of the other prokaryotic histone-like proteins and also to eukaryotic histories H 2 A and H 2 B. The protein precipitated upon heating, which is in contrast to the heat-stable feature of the other histone-like proteins. Furthermore, DNA and RNA syntheses in vitro were not affected by the presence of the protein. In view of these characteristics, this protein may play a role in maintaining the bacterial nucleoid structure.View full abstractDownload PDF (741K) -
Shio MAKINO, Hiroshi NAKASHIMA1982 Volume 92 Issue 4 Pages 1069-1077
Published: October 01, 1982
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSBovine band 3 which had been cleaved into two membrane intercalated fragments by extracellular chymotryptic digestion was partially purified and the behavior of fragmented band 3 was compared with that of intact band 3 in nonionic detergent, C12E9, solution. The two fragments were always co-eluted with nearly a 1:1 molar ratio from ionexchange and gel filtration columns, confirming the observation of Reithmeier ((1979) J. Biol. Chem. 254, 3054-3060) that the fragments are bound together. However, as shown by the molecular size of fragmented band 3 and its reactivity with dimethylmaleic anhydride and/or a cross-linking reagent, the frag-mentation caused some change in gross conformation of the protein. Dissociation of the two fragments, though not complete, was found to occur on treatment of fragmented band 3 with dimethylmaleic anhydride, which is a dissociating reagent for band 3 oligomers into the monomers. The disruption of self-associated band 3 was accompanied with a disruption of helical structure. The results suggest that a mutual interaction between the chymotryptic fragments is of importance to form the folded structure maintaining a band 3 dimer.View full abstractDownload PDF (651K) -
Koji KURODA, Michiyo HIROSE, Mitsuko OKADA1982 Volume 92 Issue 4 Pages 1079-1085
Published: October 01, 1982
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSCytosolic aspartate aminotransferase from rat liver was separated into at least 3 subforms, focused at pH 6.2, 5.9, and 5.7, by isoelectric focusing. Increase of subforms with low pI values was observed in pyridoxine-deficient rat liver. These subforms with low pI values showed low catalytic activities relative to their antigenic activities. The Km values for substrate and optimal pH values of the two main subforms were not significantly different in pyridoxine-deficient and control rat livers. Some conformational change of enzyme in the cytosol of pyridoxine-defi-cient rat liver was suggested by circular dichroic and fluorescent spectra. The N and C terminals of the enzyme from both pyridoxine-deficient rats and controls were shown to be alanine and glutamine, respectively.View full abstractDownload PDF (990K) -
Takashi KONDO, Kazue NAGATA, Mayumi SHIBUYA, Mitsuko OKADA1982 Volume 92 Issue 4 Pages 1087-1091
Published: October 01, 1982
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSThe rates of synthesis of aspartate aminotransferase isozymes in the liver and skeletal muscle in pyridoxine-deficient rats were examined. The rates of synthesis were compared in rats given pyridoxine-deficient diet ad libitum, rats given control diet ad libitum and rats pair-fed with those on the deficient diet. The rates of incor-poration of 3H-L-leucine by both cytosolic and mitochondrial enzymes were highest in pair-fed controls. Incorporation of radioactivity into the cytosolic enzyme was higher in deficient rat liver than in that of controls fed ad libitum, but the rate of incorporation into the mitochondrial enzyme was similar in these two groups. In muscle the rates of incorporation of labeled leucine into both isozymes were similar in all groups when expressed relative to total protein synthesis. It was suggested that increase of glucocorticoid receptor might result in increased synthesis of cytosolic aspartate aminotransferase in pyridoxine-deficient rat liver.View full abstractDownload PDF (380K) -
Shuji KAWATA, Takayuki IMAMURA, Kazuto NINOMIYA, Satoru MAKISUMI1982 Volume 92 Issue 4 Pages 1093-1101
Published: October 01, 1982
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSAn aminopeptidase was purified about 700-fold from porcine liver homogenate by ammonium sulfate fractionation and a series of column chromatographies on DEAE-cellulose, Sephadex G-150, DEAE-Sepharose, and hydroxyapatite. The purified enzyme had a specific activity of 4.6μmol•min-1•mg-1 using L-leucine β-naphthylamide as the substrate. The molecular weight of the enzyme was about 96, 000 as determined by Sephadex G-150 column chromatography. The enzyme had a' broad specificity and a pH optimum between 6.5 and 7.0 for hydrolysis of α-aminoacyl-β-naphthylamines, and it hydrolyzed the β-naphthylamides of aliphatic, basic, and aromatic amino acids. The enzyme also hydrolyzed peptide substrates with phenylalanine residues as their amino-termini, but it did not hydro-lyze L-phenylalanyl-L-proline. The enzyme was inhibited by metal-chelating agents, sulfhydryl reagents, heavy metals, bestatin, and puromycin. Activity of the enzyme inhibited by sulfhydryl-reactive reagents was restored by the addition of sulfhydryl compounds. The enzyme was activated by cobaltous ion and the values of both Km and Vmax increased. The activation was pH-dependent and above pH 7.5 cobalt ion behaved as an inhibitor of the enzyme. No metal ions other than cobaltous ion stimulated the enzyme appreciably.View full abstractDownload PDF (1148K) -
Makoto KOSHITA, Mariko YAMAMOTO, Ken HOTTA1982 Volume 92 Issue 4 Pages 1103-1108
Published: October 01, 1982
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSFragmented sarcoplasmic reticulum (FSR) of bullfrog skeletal muscle was frac-tionated into light and heavy sarcoplasmic reticulum (LSR and HSR) by sucrose density gradient centrifugation. Morphological and biochemical studies revealed that large parts of LSR and HSR were derived from longitudinal reticulum and terminal cisternae of SR, respectively. The Ca2+ uptake ability and ATPase activity of LSR were higher than those of HSR. Ca2+ release from Ca2+ preloaded SR vesicles by changing the medium from K-gluconate to KCl was suppressed by addition of 0.3M sucrose or glucose; there was no correlation between Ca2+ release and membrane potential change either in LSR or HSR vesicles. Dantrolene sodium (DAN, 20μM) had no effect on Ca2+ release. It is concluded that ion-induced Ca2+ release from SR (both HSR and LSR) in the isolated system is due to an osmotic effect.View full abstractDownload PDF (455K) -
Toshiaki IMAGAWA, Sabu KASAI, Kunio MATSUI1982 Volume 92 Issue 4 Pages 1109-1121
Published: October 01, 1982
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSA new peroxide compound (ML-X) was isolated from an autoxidation product of methyl linoleate and was determined as methyl 9-hydroperoxy-12, 13-epoxy-10-octadecenoate. This compound inhibited state 3 respiration of rat heart- and liver mitochondria when glutamate and malate were used as substrates, but not when the substrate was succinate. State 4 respiration of mitochondria was not affected when glutamate-malate was used as the substrate, but it was stimulated when the substrate was succinate. ML-X inhibited oxidative phosphorylation of the mito-chondria and abolished the membrane potential formed by respiration or by added ATP. NADH oxidase activity of submitochondrial particles was inhibited by ML-X but succinate oxidase activity was not inhibited. NADH-acceptor reductase activities of submitochondrial particles were inhibited by ML-X to the same extents as by rotenone. These findings show that ML-X has dual effects on mitochondrial respiration as (1) an inhibitor of NADH dehydrogenase complex and (2) an uncoupler. Neither methyl linoleate monohydroperoxide nor methyl epoxy stearate has such effects. ML-X is a new type of inhibitor-uncoupler of mitochondrial respiration in which hydroperoxy- and epoxy groups co-operate.View full abstractDownload PDF (990K) -
Kunio YAMATO, Akira YOSHIDA1982 Volume 92 Issue 4 Pages 1123-1127
Published: October 01, 1982
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSLactosylceramide and paragloboside were synthesized from their precursor glyco-lipids and UDP-galactose by lactose synthase A protein [UDP-Gal: GlcNAc β-4-galactosyltransferase, EC 2. 4. 1. 22] purified to homogenity from human plasma. The partially purified human liver enzyme and an extract from human lympho-blastoid cells also exhibited the above activities. Rabbit antibody against the purified human plasma lactose synthase A protein neutralized the glycolipid synthesis activity as well as the activity for lactose synthesis by the enzyme preparations from plasma, liver and lymphoblastoid cells. These results suggest that lactose synthase A protein existing in plasma, liver and lymphoblastoid cells can synthesize not only lactose but also lactosylceramide and paragloboside in vitro. The enzyme could play a role in the synthesis of these two glycolipids in vivo.View full abstractDownload PDF (1049K) -
Youko HISHIKAWA-ITOH, Isamu SUGIE, Hisao KATO, Sadaaki IWANAGA1982 Volume 92 Issue 4 Pages 1129-1140
Published: October 01, 1982
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSA factor, named SK-potentiator, which is known to potentiate the activation of plasminogen by streptokinase (SK), was isolated from human plasma. The procedures consisted of column chromatographies on DEAE-Sephadex A-50 and heparin-agarose, followed with gel-filtration on a Sepharose 6 B column and affinity chromatography on a lysine-Sepharose 4 B column. The isolated SK-potentiator markedly potentiated the rate of activation of plasminogen by streptokinase. However, it did not show a potentiating effect on the activation of plasminogen by urokinase or on the plasmin activity. SK-potentiator showed a similar mobility to that of fibrinogen on both SDS-polyacrylamide gel and agarose gel electrophoresis, and cross-reacted with anti-fibrinogen antiserum. The amino acid composition of SK-potentiator was very close to that of human fibrinogen, although the content of serine and threonine was significantly lower. SK-potentiator showed a single band with a molecular weight of 300, 000 on SDS-gel electrophoresis in the absence of 2-mercaptoethanol. In the presence of 2-mercaptoethanol, it showed two major bands with molecular weights of 53, 000 and 48, 000, respectively, which corre-sponded to the Bβ chain and γ chain of fibrinogen.
To establish further that the isolated SK-potentiator may be one of the fibrinogen degradation products (FDP), human fibrinogen was digested with plasmin and the SK-potentiator activity generated in the course of the digestion was measured. As a result, the SK-potentiator activity was found to initially increase and thenView full abstractDownload PDF (1972K) -
Takayoshi IIO, Hiroshi KONDO1982 Volume 92 Issue 4 Pages 1141-1149
Published: October 01, 1982
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSIncorporation of skeletal muscle troponin C (TN-C) subunit into skeletal muscle troponin (TN) induces a large increase in the apparent binding constant of Ca2+ to the low affinity Ca2+-binding sites of TN-C (from 1×105M-1 to 5.6×106 M-1 in the presence of 2mM MgCl2), and a large decrease in the rate constant of the Ca2+ removal reaction from the low affinity Ca2+-binding sites of TN-C (from 230s-1 to 37 s-1 in the presence of 2mM MgCl2). On the other hand, no significant modification in the molecular kinetic mechanism of the local conformational change due to the Ca2+ binding or removal reaction with the high affinity Ca2+-binding sites of TN-C is observed as TN-C is incorporated into TN.View full abstractDownload PDF (697K) -
Kazuko SHIBATA-SEKIYA1982 Volume 92 Issue 4 Pages 1151-1162
Published: October 01, 1982
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSTo determine whether or not the two heads of myosin from striated adductor muscles of scallop are nonidentical and the main intermediate of the ATPase reaction, MADPP, is produced only on one of the two heads, the Pi-burst size, the amount of total bound nucleotides and the amount of bound ADP during the ATPase reaction were measured in this study. The Pi-burst size was 1 mol per mol in the presence of 0.1-5mM Mg2+ ions. The amount of total nucleotides bound to myosin was 2 mol per mol. Both the amounts of bound ADP and ATP at sufficiently high ATP concentrations were 1 mol per mol of striated adductor myosin, and the affinity for ADP binding was higher than that for ATP binding. These findings indicate that MADPP or MATP is produced on each of the two heads of striated adductor myosin on its interaction with ATP.
The fluorescence intensity at 340 nm of striated adductor myosin was enhanced by about 7% upon addition of ATP. The time for the half maximum fluorescence enhancement, τ1/2, at 5μM ATP was 0.25s, which was almost equal to the τ1/2 values for the Pi-burst and for the dissociation of actomyosin reconstituted from striated adductor myosin and skeletal muscle F-actin. The dependences on ATP concentration of the extent of the fluorescence enhancement and the dissociation of actomyosin could be explained by assuming that these changes are associated with the formation of MPDP on one of the two heads of myosin.
The Pi-burst size and the amount of bound ADP of smooth adductor myosin were slightly but significantly larger than 1 mol per mol. Both ATPase reactions of striated and smooth adductor myofibrils showed the substrate inhibition. The extent of substrate inhibition of ATPase of smooth adductor myofibrils was less than that of striated adductor myofibrils. All the present findings support the view that the nonidentical two-headed structure is required for substrate inhibition of the actomyosin ATPase reaction.View full abstractDownload PDF (852K) -
Kaoru SHINOMIYA, Takaaki SHINOMIYA, Ken-ichi YOSHIDA, Hiroshi KIMURA1982 Volume 92 Issue 4 Pages 1163-1171
Published: October 01, 1982
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSWe have studied the altered electrophoretic mobility from the α2-globulin region to the α1-globulin region of the group-specific component (Gc protein) (vitamin D binding protein) in plasma of aged blood, plasma from cadavers or extracts of blood stains by agar-gel immunoelectrophoresis and polyacrylamide gel electro-phoresis. The alteration of migration of Gc protein found in plasma of aged blood is reproduced by incubation of plasma of fresh blood with extracts of platelets. Platelet extracts had no effect on the electrophoretic mobility of 20 other serum proteins examined. The alteration of electrophoretic mobility of Gc protein was not due to digestion by proteolytic enzymes or neuraminidase since polyacrylamide gradient gel electrophoresis and gel filtration studies revealed a higher molecular weight for altered Gc protein (Mr 102, 000) compared to native Gc protein (Mr 60, 000). A factor found in platelet extracts was nondialyzable, heat-labile and sensitive to trypsin and chymotrypsin, and was present in all tissues of human and rat examined. The results indicate that the alteration of the electrophoretic mobility of Gc protein found in plasma of aged blood is due to binding of a protein factor released from platelets and other blood cells during aging. The protein factor was identified as actin by partial purification of the factor from platelet extracts. Polyacrylamide gradient gel electrophoresis of fractions containing the factor showed an actin band, which comigrated with purified actin from chicken gizzard. The actin band disappeared on addition of purified Gc protein and a new protein band of Mr 102, 000 appeared. The band of Mr 102, 000 disappeared with the appearance of another new protein band of Mr 132, 000 on further addition of DNase I, which is known to bind actin.View full abstractDownload PDF (2762K) -
Keiichi TAKEISHI, Osamu GOTOH1982 Volume 92 Issue 4 Pages 1173-1177
Published: October 01, 1982
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSNucleotide sequence homology among 4.5 S RNAs from various organisms was examined by computer analysis to evaluate their sequence relationships. Chloroplast 4.5 S rRNAs of wheat and tobacco were not significantly related to Escherichia coli 4.5 S RNA, but were closely related to the 3'-terminus of bacterial 23S rRNA. Significant sequence homology was found between rat Novikoff hepatoma 4.5 S RNA, and mouse and hamster 4.5 S RNAs, suggesting that these RNAs are products of a family of genes with diverged sequences. E. coli 4.5 S RNA had no significant sequence homology with any rodent 4.5 S RNAs as a whole sequence. The E. coli, mouse and hamster 4.5 S RNAs, however, were found to share a homologous 14-nucleotide sequence at the center of the molecules, which is known to exist as a conserved sequence in both Alu and Alu-equivalent sequences of mammalian DNAs.View full abstractDownload PDF (333K) -
Takashi KAGEYAMA, Kenji TAKAHASHI1982 Volume 92 Issue 4 Pages 1179-1188
Published: October 01, 1982
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSWhen Japanese monkey pepsinogen was activated at pH 2.0 in the absence of pepstatin, the activation segment of the amino (N)-terminal 47 residues was released as a single intact polypeptide. This clearly shows that the pepsinogen was activated to pepsin directly. This direct activation was called a ‘one-step’ process. On the other hand, when pepsinogen was activated at pH 2.0 in the presence of pepstatin, an appreciable amount of pepsinogen was converted to an intermediate form between pepsinogen and pepsin, although a part of pepsinogen was activated directly to pepsin. The intermediate form was generated by releasing the N-terminal 25 residues of pepsinogen. This activation through the intermediate form is thought to be a ‘two-step’ or ‘stepwise-activating’ process involving a bimolecular reaction between pepstatin-bound pepsinogen and free pepsin.View full abstractDownload PDF (2709K) -
Matsuko MORIYASU, Akio ITO, Tsuneo OMURA1982 Volume 92 Issue 4 Pages 1189-1195
Published: October 01, 1982
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSArylsulfatase C [EC 3. 1. 6. 1] was solubilized from rat liver microsomes with Triton X-100 and purified about 2, 000-fold with an overall yield of 30-40%. The purification procedure included ion-exchange chromatography, hydrophobic affinity chromatography, and gel filtration. The purified enzyme was homogeneous as judged by polyacrylamide gel electrophoresis in the presence of SDS, and its monomeric molecular weight was estimated to be about 72, 000 daltons. The molecular weight of the native enzyme was about 280, 000 daltons as determined by gel filtration in the presence of Triton X-100, suggesting a tetrameric structure for the enzyme molecule. The enzyme showed an isoelectric point of pH 8.1. From its strong affinity toward concanavalin A-Sepharose and colorimetric determination of neutral sugars by the phenol-sulfuric acid method, arylsulfatase C was identified as a glyco-protein. Analysis of the carbohydrates by gas-liquid chromatography demonstrated that the carbohydrate chains of arylsulfatase C were rich in mannose and N-acetyl-glucosamine, suggesting that they are the high mannose-type. This conclusion was supported by the results of digestion of the enzyme with endoglycosidase H.View full abstractDownload PDF (1890K) -
Matsuko MORIYASU, Akio ITO1982 Volume 92 Issue 4 Pages 1197-1204
Published: October 01, 1982
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSThe intramembranous disposition of arylsulfatase C [EC 3. 1. 6. 1] was studied. The lack of stimulation by Triton X-100 of microsomal arylsulfatase C activity indicated the outside location of the active site of the enzyme in microsomal vesicles. The exposure of arylsulfatase C on the surface of microsomal vesicles was also suggested by the binding of antibodies against the purified enzyme to intact microsomes. However, larger amounts of the antibodies were bound to microsomes in the presence of a low concentration of Triton X-100, suggesting the presence of other antigenic sites of the enzyme not available to the antibodies in intact microsomes. The treatment of solubilized and microsome-bound arylsulfatase C with transglutaminase indicated two susceptible glutamine residues per subunit of the enzyme molecule. One of the glutamine residues was labeled with transglutaminase in intact microsomes, whereas the other one became available to transglutaminase only after the addition of Triton X-100 to microsomes. These observations suggested that endoglycosidase H-sensitive carbohydrate chains of arylsulfatase C are located in the lumen of microsomal vesicles. We conclude that microsomal arylsulfatase C is a transmembranous protein and exposed on both outer and inner surfaces of the membrane.View full abstractDownload PDF (1386K) -
Itsuro NAKANO, Motoaki ANAI1982 Volume 92 Issue 4 Pages 1205-1212
Published: October 01, 1982
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSATP-dependent deoxyribonuclease from Micrococcus luteus was purified to near homogeneity by a procedure involving gentle cell lysis, ammonium sulfate fractionation, TEAE-cellulose chromatography, Sephadex G-150 gel filtration and DNA-cellulose chromatography. Treatment of the enzyme with 2, 3-butanedione, which binds specifically to arginyl residues, caused rapid loss of enzyme activities and the effect was enhanced by borate ion. The reaction obeyed first order kinetics with respect to the butanedione concentration, indicating that at least one functional arginyl residue is involved in the inactivation reaction. The enzyme was protected from inactivation by the presence of a low concentration of ATP, but not of ADP, AMP or adenosine. These results indicate that ATP-dependent deoxyribonuclease of Micrococcus luteus has functional agrinyl residue (s) at an ATP-binding site.View full abstractDownload PDF (570K) -
Hideo MAEDA, Nobuhisa NAKAMURA, Haruo UZAWA1982 Volume 92 Issue 4 Pages 1213-1218
Published: October 01, 1982
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSWe studied the proteolytic action in vitro of free and α2-macroglobulin-bound porcine pancreatic elastase [EC 3. 4. 21. 11] on the apolipoproteins of plasma: very low density lipoprotein (VLDL), low density lipoprotein (LDL), and high density lipoprotein (HDL). Polyacrylamide gel electrophoresis, isoelectric focusing and immunodiffusion tests of elastase-treated plasma lipoproteins revealed that apolipoprotein C-II and C-III polypeptides were more susceptible to elastase in free form than plasma apolipoproteins (A-I, A-II, B, and E). Elastase bound to α2-macroglobulin did not show any such activities.View full abstractDownload PDF (2815K) -
Ken-Ichi FURUKAWA, Yuji TONOMURA1982 Volume 92 Issue 4 Pages 1219-1225
Published: October 01, 1982
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSSH-1 thiol of S-1 was modified with N-(1-oxyl-2, 2, 6, 6-tetramethyl-4-piper dinyl) iodo-acetoamide spin label (IASL). The extent of dissociation, α, of spin-labeled myosin subfragment-1 (IASL-S-1) from acto-IASL-S-1 by a nucleotide was measured by an ultracentrifugal separation method, a light-scattering method, and a saturation transfer EPR method. The α values obtained by these three methods were the same within the limits of the experimental errors. The dependence of α on the concentrations of AMPPNP, [S], and F-actin, [A], could be described by the equation: α-1=1+(1+Ks/[S])[A]/KA. The Ks and KA values were 0.65-1.2mM and 1.7-2.7mg/ml, respectively, in 0.5M KCl and 4mM MgCl2 at pH 7.0 and 20°C. The height of the weakly immobilized peak of the conventional EPR spectrum of IASL-S-1, W, increased linearly with increase in the ATP or AMPPNP concentration, and became saturated at 1 mol nucleotide/mol IASL-S-1. No change in W was observed upon the binding of IASL-S-1 with F-actin. The dependence of the extent of change in W, ΔW, on [A] and [S] was given by ΔW-1=1+-Ks/[S], where -Ks=Ks/(1+KA/[A]). This finding indicates that the ΔW value is proportional to the amount of a nucleotide bound to IASL-S-1 and independent of the binding of F-actin to IASL-S-1.View full abstractDownload PDF (517K) -
Yasuko KAWAMURA, Hideyo HASUMI, Satoshi NAKAMURA1982 Volume 92 Issue 4 Pages 1227-1233
Published: October 01, 1982
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSThe reconstitution reaction of deoxy hemoglobin (Hb) tetramer from isolated α and β chains was kinetically studied by measuring circular dichroism (CD) changes in the Soret and the ultraviolet regions and optical absorbance change in the Soret region with a stopped-flow apparatus. The CD change in the Soret region was a fast reaction, followed by that in the ultraviolet region and the absorbance change in the Soret region. This fast reaction followed a simple second-order rate law with a rate constant of 6.4×105M-1•s-1. These results indicated that the combination of α and β monomers into an αβ dimer accompanied the CD change in the Soret region and was the rate-limiting step of the overall reconstitution reaction of deoxyHb tetramer.
On the other hand, the CD change in the ultraviolet region was ascribed to the combination of two αβ dimers into an Hb tetramer. The absorbance change in the Soret region was related to both the combination of α and β monomers and that of two αβ, dimers. From the analyses of these reactions the rate constant of the combination of two αβ dimers was determined to be 1.0×106M-1•s-1.View full abstractDownload PDF (514K) -
Hirokazu KOYAMA1982 Volume 92 Issue 4 Pages 1235-1240
Published: October 01, 1982
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSL-Phenylalanine oxidase from Pseudomonas sp. P-501 has been purified to homogeneity as judged by acrylamide gel electrophoresis and ultracentrifugation. The enzyme produced both β-phenylpyruvate and α-phenylacetamide from L-phenyl-alanine.
Balance studies demonstrated that consumption of 1 mol each of L-phenyl-alanine and oxygen resulted in the formation of 0.2 mol each of β-phenylpyruvate, ammonia, and hydrogen peroxide and 0.8 mol each of α-phenylacetamide and carbon dioxide under aerobic conditions. Thus, the same enzyme preparation catalyzed simultaneous oxidative deamination and oxygenative decarboxylation of L-phenyl-alanine.
Besides L-phenylalanine, the enzyme oxidized L-tyrosine, L-methionine, and L-tryptophan at lower reaction rates.View full abstractDownload PDF (1141K) -
Akira SAKAI, Yoshio WATANABE1982 Volume 92 Issue 4 Pages 1241-1249
Published: October 01, 1982
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSDNA polymerase A (an α-type polymerise) from the ciliate, Tetrahymena pyriformis, has been purified 260, 000-fold (40, 000 units/mg protein). The polymerase A did not show any heterogeneity in terms of size and charge during purification. Enzymatic properties of the DNA polymerase A remained unchanged during the purification. Two-dimensional gel electrophoresis revealed that in the first dimension (isoelectric focusing agarose gel), the activity of the purified enzyme was focused at around pH 5.5 and that in the second dimension (sodium dodecyl sulfate-polyacrylamide gel), 135, 000- and 66, 000-dalton polypeptides emerged from the activity peak at a stoi-chiometric ratio of about 1:3. The native molecular weight of the DNA polymerase A estimated from the stoichiometric subunit ratio approximately coincided with that estimated from gel filtration on Sephadex G-200 under low ionic strength conditions.
The present results strongly suggest the existence of a common high-molecular-weight catalytic core subunit in α-type polymerases of eukaryotes.View full abstractDownload PDF (1430K) -
Hideko SUYA, Yukichi ABE, Isao TANAKA, Shin-ichi ISHII, Koichi ITAYA1982 Volume 92 Issue 4 Pages 1251-1257
Published: October 01, 1982
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSInsulin-like action of monovalent monomeric concanavalin A (m-Con A) was examined in rat adipocytes in the presence of anti-m-Con A antiserum. The antisera from rabbits injected with m-Con A reacted with not only monovalent monomeric but also tetravalent tetrameric concanavalin A (α-Con A) in Ouchterlony double diffusion analysis. m-Con A alone did not show any appreciable effect on glucose oxidation of adipocytes while it slightly inhibited glycerol release stimulated by epinephrine. In contrast, exposure of adipocytes to m-Con A in the presence of antibodies to m-Con A resulted in stimulation of glucose oxidation and inhibition of epinephrine-stimulated lipolysis. The stimulation and the inhibition with m-Con A in the presence of the antibodies were of the same degree as those with α-Con A. Both α- and m-Con A were slightly active in inhibiting 125I-labeled insulin binding. These results demonstrate that the ability of anti-m-Con A antiserum to aggregate m-Con A bound to receptors on the isolated-adipocyte plasma membrane allowed m-Con A to mimic the biological activity of insulin and that the aggregation of receptors for ligands other than insulin can induce insulin-like action in rat fat cells.View full abstractDownload PDF (557K) -
Ryoko TAMURA, Yoshiki TAKESUE, Yoshimi NISHI1982 Volume 92 Issue 4 Pages 1259-1268
Published: October 01, 1982
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSActin and four major associated proteins have been identified in the microvillus cytoskeleton from chicken intestinal brush borders isolated in the presence of Ca2+ chelating agents. We have isolated microvilli from rabbit small intestine in the presence of Ca2+ and found by sodium dodecyl sulfate-gel electrophoresis that a protein of 33, 000 daltons exists in a molar ratio to actin of 1:3 within the isolated microvillus. The protein could be extracted from acetone powder of microvilli in the absence, but not in the presence, of Ca2+. Thus, for its purification the acetone powder was extracted twice with buffer containing Ca2+ and then with buffer containing a Ca2+-chelator. The 33, 000-dalton protein was purified from the last extract by Sephadex G-100 gel filtration and DEAE-cellulose column chromatog-raphy. The purified protein was a water-soluble monomeric protein with a molecular weight of 33, 000. Antisera against the purified protein raised in guinea pigs reacted with the antigen but not with actin or tropomyosin purified from rabbit skeletal muscle. Freezing-thawing resulted in aggregation of the purified protein, and the aggregates showed filamentous and sheet-like structures. These results suggests that the 33, 000-dalton protein is a major component of the microvillus cytoskeleton.View full abstractDownload PDF (4716K) -
The Effect of Crosslinking of Thin Filament with Glutaraldehyde on the Contractility of Muscle FiberEwa PRóCHNIEWICZ-NAKAYAMA, Toshio YANAGIDA1982 Volume 92 Issue 4 Pages 1269-1277
Published: October 01, 1982
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSThe effect of crosslinking of F-actin with glutaraldehyde on the contractility of muscle ghost fiber containing reconstituted thin filament (i.e. F-actin-tropomyosin-troponin complex) and irrigated with myosin was investigated. The results show that: (i) crosslinking inhibited development of isometric tension and shortening of the fiber in the presence of MgATP, (ii) superprecipitation of the complex of myosin with crosslinked thin filament was considerably delayed, (iii) crosslinking inhibited neither binding of myosin heads to the filament nor activation of myosin ATPase. It is suggested that alterations of actin structure due to the formation of intra- and/or intermonomer crosslinks can essentially affect the process of contractility.View full abstractDownload PDF (1375K) -
Masao NAKAMURA, Isao YAMAZAKI1982 Volume 92 Issue 4 Pages 1279-1286
Published: October 01, 1982
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSWhen xanthine oxidase was prepared from fresh raw cow's milk in the presence of dithioerythritol, 94% of its xanthine-oxidizing activity was found as a dehydro-genase type. The enzyme was reversibly converted to an oxidase type when dithio-erythritol was removed. The conversion was ascribable to the oxidation of sulfhydryl groups of the enzyme by oxygen. The two forms of the enzyme gave the same visible spectrum, but the dehydrogenase form alone gave a characteristic difference spectrum upon addition of NAD+. NADH served as a good electron donor for the dehydrogenase form of the enzyme but not for the oxidase form. When xanthine was used as an electron donor, the overall rate of p-benzoquinone reduction was the same for the oxidase and dehydrogenase forms, but the proportion of one-electron flux from the enzyme to p-benzoquinone was considerably greater in the reaction of the dehydrogenase form than in that of the oxidase form.View full abstractDownload PDF (568K) -
Yutaka KIRINO, Hiroshi SHIMIZU1982 Volume 92 Issue 4 Pages 1287-1296
Published: October 01, 1982
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSUptake and release of Ca2+ in heavy and light fractions of fragmented sarcoplasmic reticulum (FSR) isolated from frog and rabbit skeletal muscle was studied under conditions similar to those employed in skinned muscle fiber experiments, where ATP and Mg2+ concentrations were considered to be physiological and free Ca2+ concentration was kept constant during the Ca2+ uptake and release. Ca2+ level in FSR monotonously approached a steady state level which depended only on the final experimental conditions. Heavy fractions, but not light fractions, exhibited characteristics similar to those of Ca2+-induced Ca2+ release reported in skinned fiber studies: i) the rate and steady state level of Ca2+ uptake increased with increase in free Ca2+ concentration in the reaction medium up to 10-6M. With further increase in free Ca2+ concentration, the steady state level of Ca2+ taken up decreased while the Ca2+ uptake rate increased. ii) The steady state Ca2+ level was decreased by caffeine but increased by procaine or ruthenium red. Parallel measurement of Ca2+-ATPase activity clearly showed that these drugs modify the Ca2+ efflux but hardly affect the Ca2+-pump activity. It was concluded that the Ca2+-induced Ca2+ release mechanism was in operation at as low as 10-6M free Ca2+ concentration. Treatment of FSR with 0.6M KCl did not have any significant effect.View full abstractDownload PDF (780K) -
Kensuke YONEMURA, Hiroshi MAEDA1982 Volume 92 Issue 4 Pages 1297-1303
Published: October 01, 1982
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSThe fluorescence polarization method was applied for the determination of DNase-activity using ethidium bromide-labeled DNA as a substrate. This assay was based on a decrease in fluorescence intensity or fluorescence polarization value, which reflected degraded molecular size resulting from the specific cleavage of DNA by the enzyme. DNase activity was quantitatively determined above 0.1 ng/ml of the enzyme. A decrease in fluorescence intensity or polarization value brought about by DNase was inhibited by the specific inhibitors of DNase such as EDTA, pyrophosphate, and thymidine-3', 5'-diphosphate. In addition, specificity of the change of polarization value was established using other enzymes. Since this method was simple, rapid and sensitive, it is applicable for both quantitative and qualitative assays as well as for the kinetic analysis of the enzyme.View full abstractDownload PDF (480K) -
Yoshiteru KOBAYASHI1982 Volume 92 Issue 4 Pages 1305-1318
Published: October 01, 1982
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSTubulin from either outer doublet microtubules or the central pair of microtubules in the starfish sperm flagellum was resolved into two α-subunits and one β-subunit by two-dimensional gel electrophoresis. Further fractionation of the doublet microtubules into B-tubules, A-tubules, and ‘partitions, ’ i.e. a part of the A-tubule wall shared with the B-tubule, revealed that all these fractions contained two α-subunits (α1 and α2) and one β-subunit of tubulin, the ratios of α/β being almost one. Both the B-tubule and the wall of the A-tubule exclusive of the partition consisted of α1-and α2-subunits in addition to β-subunits, while the partition was much richer in the α1-subunit than the α2-subunit. Peptide mapping after limited proteolysis of tubulin subunits from the central pair, the wall of outer doublet microtubules except the partition, and the partition, indicated that α-subunits and β-subunits from the former two sources were quite similar to each other, but distinguishable from the α-subunit (α1') and β-subunit of the partition. On the basis of these results, it can be said that at least three subspecies of α-subunit and two subspecies of β-subunit exist in ‘stable microtubules’ found in the sperm flagellum. Under the electron microscope, the partition fraction contained ribbons consisting of two and three protofilaments. These results suggest that each of the two adjacent protofilaments constituting the partition is a thread of α1'=β dimers. In the partition, there were also five constitutive polypeptides in addition to tubulin.View full abstractDownload PDF (6278K) -
Julian A. BARDEN, Paul M. G. CURMI, Cristobal G. dos REMEDIOS1982 Volume 92 Issue 4 Pages 1319-1323
Published: October 01, 1982
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSNMR and UV spectroscopy reveal that the tetravalent thorium ion interacts with actin in an identical fashion to the trivalent lanthanide ions. UV spectra and viscometry data suggest that the actin-bound Th4+ has an ionic radius of about 110 pm. Th4+ also contributes to the formation of crystalline actin tubes under appropriate conditions. The factors governing whether tubes form appear to be cationic radius and total charge on the actin monomer, rather than the valency of the ion per se.View full abstractDownload PDF (966K) -
Toshio MATSUYAMA, Kuniji TANAKA, Midori MASHIKO, Masako KANAMOTO1982 Volume 92 Issue 4 Pages 1325-1328
Published: October 01, 1982
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSThe enzyme of Aspergillus fumigates which produces di-D-fructose 1, 2'; 2, 1' dianhydride (difructose anhydride I) (Tanaka et al., 1979) was investigated for its mode of action. Fractionation and purification of the enzyme showed that it produced difructose anhydride I not from inulin but from inulobiose by the reverse reaction of the partial hydrolysis of difructose anhydride I.View full abstractDownload PDF (786K) -
Shigeharu NAGASAWA, Kiyoshi MIZUGUCHI, Chikako ICHIHARA, Jiro KOYAMA1982 Volume 92 Issue 4 Pages 1329-1332
Published: October 01, 1982
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSHuman C 4 binding protein (C 4 bp), which is a macromolecular weight (Mr 450, 000-590, 000) cofactor of C 3 b/C 4 b inactivator (I), is composed of 6 or 8 disulfide-linked polypeptide chains of Mr 75, 000. Chymotrypsin cleaved C 4 bp into two major fragments; a large fragment of Mr 160, 000, which contained carbohydrate chains and was composed of disulfide-linked polypeptide chains of Mr 25, 000, and a small fragment of Mr 48, 000, which was a single polypeptide chain and had the cofactor activity of C 4 bp. These results suggest that chymotrypsin liberates a functional domain-containing Mr 48, 000 fragment from each subunit chain of C4bp and yields a core fragment derived from a disulfide-knot domain connecting each subunit chain of C4bp.View full abstractDownload PDF (2328K) -
Yoshinori MORIYAMA, Tatsuya TAKANO, Shoji OHKUMA1982 Volume 92 Issue 4 Pages 1333-1336
Published: October 01, 1982
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSAcridine orange was found to accumulate in pure lysosomal particles (tritosomes) in vitro, and the quenching of its fluorescence correlated well with the ΔpH (inside acid) across the lysosomal membrane, Use of this dye showed that Mg-ATP caused lysosomal acidification. This acidification was sensitive to N, N'-dicyclohexylcar-bodiimide, N-ethylmaleimide, and azide, but not to oligomycin, ouabain or vanadate. These results supported the idea of the existence of a lysosomal H+-pump, suggested in a previous paper (Ohkuma et al. (1982) Proc. Natl. Acad. Sci. U. S. 79, 2758-2762).View full abstractDownload PDF (291K)
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