巻号一覧
89 巻, 4 号
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Toshiaki KATADA, Michio UI1981 年 89 巻 4 号 p. 979-990
発行日: 1981/04/01
公開日: 2008/11/18
ジャーナル フリーPancreatic islets were maintained in culture with or without islet-activating protein (TAP), which is a new protein purified from culture medium of Bordetella pertussis. These cultured islets (IAP-treated or control) were then incubated for 30min in IAP-free medium with various insulin secretagogues. During incubation, much more insulin was released from IAP-treated islets than control islets in response to glucose, arginine, glucagon, and sulfonylurea. IAP was effective in this regard when added to cultures at concentrations higher than 0.01 ng/ml; the effect was dependent on concentration up to 100 ng/ml. Enhanced insulin secretion was associated with accumulation of cyclic AMP when breakdown of the nucleotide was prevented by a methylxanthine. Epinephrine caused marked inhibitions, via α-adrenergic receptors, of glucose-induced insulin release, cyclic AMP accumulation and 45Ca uptake in control islets but did not in IAP-treated islets during incubation. None of these effects of TAP pretreatment were observed unless the medium for incubation was supplemented with Ca ions. 45Ca ion flux through the islet cell membrane was accelerated by the IAP treatment; conceivably, IAP was effective in causing sustained activation of native calcium ionophores on the membrane, which would be responsible for the enhanced insulin and cyclic AMP responses characteristic of IAP-treated islets.抄録全体を表示PDF形式でダウンロード (965K) -
Mikiharu YOSHIDA, Norio KATOH, Shuichiro KUBO, Koichi YAGI1981 年 89 巻 4 号 p. 991-997
発行日: 1981/04/01
公開日: 2008/11/18
ジャーナル フリーAntisera against the isozymes of heavy meromyosin from chicken breast muscle myosin were prepared by immunizing rabbits. Both antisera against heavy meromyosin containing gl light chain (HMM(g1)) and that containing g3 light chain (HMM(g3)) reacted with both heavy meromyosin isozymes and myosin, but not with whole light chain mixture. This indicates that only antibodies against the heavy chain of heavy meromyosin were elicited. The antisera were applied successively to two columns, one coupled with subfragment-2 and the other with heterologous subfragment-1 isozymes. When the antiserum against HMM(g3) was absorbed with subfragment-1 containing gl (S-1(g1)), it did not react with HMM(g1), but reacted with HMM(g3). On the other hand, when the antiserum against HMM(g1) was absorbed with subfragment-1 containing g3 (S-1(g3)), it lost its ability to react with both heavy meromyosin isozymes. This indicates the presence of a specific antibody against the heavy chain of HMM(g3). The head portion of myosin containing g3 may hold an unique antigenic determinant which is not present in the head of a myosin containing g1.抄録全体を表示PDF形式でダウンロード (1714K) -
Yuji SUGITA, Tokuhiko HIGASHI1981 年 89 巻 4 号 p. 999-1004
発行日: 1981/04/01
公開日: 2008/11/18
ジャーナル フリーInactive catalase [EC 1. 11. 1. 6] occurring inconsiderable amounts in liver peroxisomal extracts from Sedormid-treated rats was investigated. The antigen-to-enzyme ratio (i.e. the ratio of catalase determined immunochemically to catalase determined enzymatically; 1.0 for purified enzyme) was around 1.7 when peroxisomal extract from Sedormid-treated rats was assayed.
After dialysis of the extract against a solution containing 44mM acetate buffer, pH 4.1, and 22% ethanol, catalase remaining in soluble from showed the same antigen-to-enzyme ratio as the purified enzyme (1.0). Moreover, the total enzymatic activity was not reduced. The results indicated that the catalase precipitated by this treatment was enzymatically inactive.
The peroxisomal extract was analyzed by gel filtration, and an enzymatically inactive catalase was detected in the region corresponding to molecules smaller than the enzymatically active catalase (tetramer). When the immunoprecipitates of the inactive catalase were analyzed by polyacrylamide gel electrophoresis in the presence of SDS and urea, only a subunit (monomer) of catalase was detected. At 60min after injection of [14C] amino acid and δ-[3H] aminolevulinic acid into rats, the specific radioactivity of 14C in this inactive catalase was 6-7 times that of active catalase. On the other hand, the 3H/14C ratio was much lower than that of active catalase.
The results suggest that the inactive catalase isolated by gel filtration is an intermediate of maturation, in which apo-catalase monomer binds with heme and assembles to form complete molecules.抄録全体を表示PDF形式でダウンロード (946K) -
Masanori IWAMA, Motoko KUNIHIRO, Kazuko OHGI, Masachika IRIE1981 年 89 巻 4 号 p. 1005-1016
発行日: 1981/04/01
公開日: 2008/11/18
ジャーナル フリー1. Two RNases (RNase UL and RNase Us) were purified from the urine of human adults by means of column chromatographies on SP-Sephadex C-50, phospho-cellulose and CM-cellulose and gel-filtration on Sephadex G-75 in homogeneous states obtained by SDS-disc electrophoresis.
2. Molecular weights of these RNases determined by gel-filtration were 38, 000 and 13, 000 for RNase UL and RNase US, respectively.
3. Optimal pH's of urine RNases were 8.0 and 6.75 for RNase UL and RNase US, respectively.
4. Chemical composition of urine RNases was determined. RNase UL contains about 20.7% of neutral sugar and 7.8% of hexosamine. RNase Us contains a very small amount of carbohydrate moiety.
5. Base specificity of urine RNases studied with 2', 3'-cyclic nucleotides and dinucleoside phosphates as substrates indicated that both RNases were pyrimidine specific and cytosine preferential enzyme, as is bovine pancreatic RNase A. Although base specificity of RNase UL was qualitatively similar to RNase A, that of RNase US was slightly different. That is, RNase US did not hydrolyze UpU and hydrolyzed UpC and 2', 3'-cyclic UMP very slowly.
6. Antigenic properties of human urine RNases were studied by Ouchterlony's double diffusion analysis. RNase UL, RNase US, and RNase A were serologically distinguishable.抄録全体を表示PDF形式でダウンロード (1867K) -
Naoki KAWADA, Katsumichi TAKEDA, Yoshiaki NOSOH1981 年 89 巻 4 号 p. 1017-1027
発行日: 1981/04/01
公開日: 2008/11/18
ジャーナル フリーNADH dehydrogenase [EC 1. 6. 99. 3] in membranes of Bacillus caldotenax was solubilized with sodium N-lauroylsarcosinate and purified 50-fold from membranes to 75-80% homogeneity, as judged by SDS-polyacrylamide gel electrophoresis. The enzyme was considered to be located on the electron transport chain and to be an FAD-containing protein. The molecular weight of the subunit was estimated to be 44, 000. The enzyme (or the enzyme bound to the B. caldotenax membrane lipids) follows a ping-pong mechanism. The enzyme can oxidize NADH, but not NADPH, with 2, 6-dichlorophenol indophenol, ferricyanide, menadione, and cytochrome c as electron acceptors. Membrane lipids or Triton X-100 stimulated the enzyme activity, except that with menadione. Lipids decreased the apparent affinity of electron acceptors and NADH to the enzyme, and increased the maximum velocity, except when menadione was used as the electron acceptor. Lipids partially protected the enzyme from thermal inactivation. The enzyme exhibited a continuous Arrhenius plot, while the lipids- or membrane-bound enzyme exhibited a discontinuous plot.抄録全体を表示PDF形式でダウンロード (1416K) -
Taichi USUI, Sumiko HOSOKAWA, Takashi MIZUNO, Tateo SUZUKI, Hiroshi ME ...1981 年 89 巻 4 号 p. 1029-1037
発行日: 1981/04/01
公開日: 2008/11/18
ジャーナル フリーA heterogalactan was isolated from the hot water extract of fruit bodies of Fomitopsis pinicola by a combination of fractionation procedures including precipitation with ethanol and with Cetavlon, and chromatography on columns of DEAE-celluose and Sephadex G-100. Despite its apparent homogeneity on gel filtration, zone electrophoresis, sedimentation equilibration, and immunodiffusion analyses, the neutral component of heterogalactan was further fractionated into unbound, weakly bound, and strongly bound forms by affinity chromatography on a column of concanavalin A-Sepharose CL 4 B. The former two polysaccharides fractions eluted with 0.1M phosphate buffer (pH 7.0) were found to be a fucogalactan and a mannofucogalactan, respectively. A more tightly bound fraction (mannofucogalactan) was subsequently eluted with 0.1M glucose in 1M NaCI.
The results of methylation, complete Smith degradation, and proton and 13C NMR spectroscopic analyses indicated that the three kinds of heterogalactans are all highly branched polysaccharides containing a framework of (1→6)-linked α-D-galactopyranosyl residues, the C-2 positions of which are substituted in different proportions with either single L-fucopyranosyl residues or disaccharide units of 3-O-α-D-mannopyranosyl-L-fucopyranose residues.抄録全体を表示PDF形式でダウンロード (1174K) -
Takeshi MIZUNO1981 年 89 巻 4 号 p. 1039-1049
発行日: 1981/04/01
公開日: 2008/11/18
ジャーナル フリーA novel peptidoglycan-associated lipoprotein (PAL) was found in the cell envelope of Proteus mirabilis. This protein showed the following properties: (1) The apparent molecular weight in sodium dodecyl sulfate (SDS) polyacrylamide gel was 18, 000. (2) The protein was present in the cell envelope in a form very closely, but not covalently, associated with the peptidoglycan layer. (3) The protein was recovered predominantly from the outer membrane fraction after separation of the cell envelopes. (4) [1 14C] Palmitic acid and [2-3H] glycerol were incorporated into the protein. (5) The protein contained covalently linked fatty acids (about 3 mol of fatty acid per mol of protein). (6) An unidentified compound was present in the hydrolysate of the protein. These properties, except for molecular weight and non-covalent association with the peptidoglycan, showed resemblance to those of Braun's lipoprotein. However the protein was distinct from Braun's lipoprotein in regard to amino acid composition. A similar peptidoglycan-associated lipoprotein (PAL) was present widely in the cell envelopes of various Gram-negative bacteria. P. mirabilis contains about twelve times as much PAL as Escherichia coli. Antiserum against PAL of P. mirabilis was cross-reactive against PAL of E. toll, but not against Braun's lipoprotein of E. coll.抄録全体を表示PDF形式でダウンロード (3167K) -
Takeshi MIZUNO1981 年 89 巻 4 号 p. 1051-1058
発行日: 1981/04/01
公開日: 2008/11/18
ジャーナル フリーIn the outer membrane of P. inirabilis, a peptidoglycan-associated lipoprotein (PAL) of apparent molecular weight 18, 000 is present as a major protein. A fatty acidcontaining polypeptide (lipopolypeptide; LPP) was isolated by digestion of the purified PAL with trypsin in the presence of 0.05% SDS. It was composed of 31 amino acid residues, an unidentified compound [X], and ca. 3 fatty acid residues. A lipooligopeptide (LOP) was also isolated after further digestion of LPP with trypsin in the absence of SDS. LOP was composed of 4 amino acid residues (Asx, 2Ser, Lys), a compound [X], and ca. 3 fatty acid residues. The C-terminal amino acids of LPP and LOP were determined as arginine and lysine, respectively. On the other hand, the N-terminus of PAL, LPP, or LOP could not be identified by conventional N-terminal analysis, indicating that the N-terminus is probably masked. These results indicate that LPP and LOP are derived from the fatty acid-attached amino terminal region of PAL.抄録全体を表示PDF形式でダウンロード (1271K) -
Takeshi MIZUNO1981 年 89 巻 4 号 p. 1059-1066
発行日: 1981/04/01
公開日: 2008/11/18
ジャーナル フリーThe amino acid sequence of the lipooligopeptide (LOP) derived from the N-terminal region of the peptidoglycan-associated lipoprotein (PAL) of Proteus mirabilis was determined to be [X]-Ser-Ser-Asn-Lys. An unidentified compound [X] present at the N-terminus was identified as glycerylcysteine [S-(propane-2', 3'-diol)-3-thio, 2-amino-propanoic acid]. The partial amino acid sequence of the lipopolypeptide (LPP), which contained the lipooligopeptide (LOP) at its N-terminal part, was also determined, mainly by Edman-degradation. The structure of the N-terminal part of PAL was determined to be [3 Fatty acids_??_glycerylcysteine-Ser-Ser-Asn-Lys-Asn-Asp-Asp-Glu-Thr-Asp-Thr-Ser••••]. The structure of PAL is discussed in comparison with Braun's lipoprotein.抄録全体を表示PDF形式でダウンロード (563K) -
Mariko IWASAKI, Sadako INOUE1981 年 89 巻 4 号 p. 1067-1074
発行日: 1981/04/01
公開日: 2008/11/18
ジャーナル フリーOne of the major fractions of sialoglycoprotein isolated from herring eggs has been shown to contain both alkali-labile and alkali-stable carbohydrate units.
Homogeneous preparations of reduced disaccharide and trisaccharide were obtained. Their structures, as determined by methylation analysis coupled with GLC-mass spectrometry, NMR spectroscopy and digestion with glycosidases, were β-n-galactopyranosyl-(1→3)-N-acetyl-D-galactosaminitol and β-D-galactopyranosyl-(1→3)-[α-N-acetylneuraminyl-(2→6)]-N-acetyl-D-galactosaminitol, respectively. No other oligosaccharide was found among the products of alkaline borohydride treatment.
Glycopeptide fractions were obtained from the pronase digest of the glycoprotein. The carbohydrate components of small glycopeptide fractions were either galactose and N-acetylgalactosamine or galactose, N-acetylgalactosamine and N-ace-tylneuraminic acid and accounted for 80% of reduced oligosaccharides obtained by alkaline borohydride treatment of the glycoprotein. Threonine was the only amino acid that occurred in molar ratios relative to N-acetylgalactosamine of more than unity in these small glycopeptide fractions. It was concluded that, on average, 4 carbohydrate units are attached to each peptide chain through O-glycosidic linkages between the N-acetylgalactosamine and threonine residues.抄録全体を表示PDF形式でダウンロード (576K) -
Takashi SHIMAKATA, Takashi KUSAKA1981 年 89 巻 4 号 p. 1075-1080
発行日: 1981/04/01
公開日: 2008/11/18
ジャーナル フリー2-Enoyl-CoA reductase was purified to homogeneity for the first time from the crude extract of Mycobacterium smegmatis. Its molecular weight was estimated to be 26, 000 by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. NADH acted as an electron donor for the reduction of 2-enoyl-CoA, while NADPH did not. The Km value for NADH was 21.3 μm. On the other hand, NAD inhibited the reaction by competing against NADH, and the K1 value for NAD was 47 μm. Among the enoyl-CoAs used as substrates, those having C10-C161 were found to be most suitable substrates for the purified reductase in terms of both apparent Km and Vmax values. The enzyme was strongly inhibited, however, when the concentration of the C16-substrate was over 50 μm, The enzyme had almost no activity towards substrates having less than C8. When NAD3H was used as an electron donor to 2-dodecenoyl-CoA in the presence of the purified reductase, only laurate was tritiated as the product.
Diacetyl and phenylglyoxal, agents that react specifically with arginine, inactivated the reductase in a time- and concentration-dependent manner during the preincubation. These results suggest that some arginine residues in the reductase protein are involved in the enzyme activity.抄録全体を表示PDF形式でダウンロード (953K) -
Hideo NAKAGAWA, Masayuki ISAJI, Masahiko HAYASHI, Susumu TSURUFUJI1981 年 89 巻 4 号 p. 1081-1090
発行日: 1981/04/01
公開日: 2008/11/18
ジャーナル フリーA rapid breadkown of collagen was found in granulation tissue induced by carrageenin in rats; the half-life of collagen in both growing and resorbing tissues was about 3.5 days, whereas that in non-resorbing tissue was about 7 days. On the other hand, the half-life of noncollagen protein in the growing, resorbing and nonresorbing tissues was about 2-3 days.
ε-Amino-n-caproic acid n-hexyl ester, an inhibitor of plasmin and trypsin, selectively inhibited collagen breakdown in vivo without affecting the degradation of noncollagen protein or the syntheses of collagen and noncollagen protein in granulation tissues. A similar selective inhibition of collagen breakdown was also found upon treatment with soybean trypsin inhibitor. Collagenase activity was assayed directly in the insoluble 6, 000×g pellet of granulation tissue homogenates. ε-Amino-n-caproic acid n-hexyl ester and soybean trypsin inhibitor markedly inhibited the collagen breakdown in granulation tissue pellets in vitro. The results are consistent with those from in vivo experiments and suggest that both the inhibitors indirectly inhibit the collagen breakdown in granulation tissue through the inhibition of a latent collagenase-activating proteinase (s), because none of the inhibitors directly inhibit collagenase.
It may be argued, therefore, that a proteinase (s) which activates a latent collagenase plays an important role in the rapid breakdown of collagen in granulation tissues.抄録全体を表示PDF形式でダウンロード (689K) -
Susumu IWASA, Chieko KITADA, Isamu YOSHIDA, Koichi KONDO, Masatake HOR ...1981 年 89 巻 4 号 p. 1091-1099
発行日: 1981/04/01
公開日: 2008/11/18
ジャーナル フリーA highly specific enzyme immunoassay for determining hCG was established by using β-D-galactosidase as label. In order to increase the specificity of the assay, an antiserum against whole hCG was purified on a column of hCG β carboxylterminal peptide (residues 123-145) covalently linked to Sepharose 4 B. The antibody (N101S) thus prepared showed a weak cross-reactivity with human LH in an assay using hCG-enzyme conjugate, but the slight cross-reactivity was virtually avoided when an hCG β carboxyl-terminal peptide was used as a peptide in the enzyme conjugate.
N101S antibody was compared with antiserum (BIB) directed against a carboxyl-terminal peptide (123-145). In hCG measurement N101S gave about 30 times higher sensitivity than BIB, although the former antibody was less sensitive to carboxyl-terminal peptides of hCG β. The enzyme immunoassay using a combination of N101S antibody and a carboxyl-terminal peptide (130-145)-enzyme conjugate was able to detect as little as 0.25 mIU of hCG without the interference of LH. The performance and validity of this assay were comparable to those of conventional radioimmunoassay.抄録全体を表示PDF形式でダウンロード (635K) -
Ken-ichi IIJIMA, Jun-ichi KISHI, Taro HAYAKAWA1981 年 89 巻 4 号 p. 1101-1106
発行日: 1981/04/01
公開日: 2008/11/18
ジャーナル フリー1. Active type collagenase was purified as much as 140-fold from the explant medium of bovine dental sacs and showed a single band on disc gel electrophoresis. Purified collagenase cleaved native collagen at only one locus under physiological conditions, but hydrolyzed neither gelatin nor α-casein. The optimal pH was about 7.8.
2. The molecular weight of active type enzyme was 35, 000 by gel filtration and 34, 000 by gel electrophoresis. The activation of latent type collagenase resulted in the reduction of molecular weight from 45, 000 to 38, 000 by gel filtration.
3. A small but detectable amount of collagenase was directly extracted from frozen and thawed bovine dental sacs. In explant media of frozen and thawed tissue and fresh tissue with actinomycin D, some activity was detected for the first 2 days, but essentially no collagenase activity was detected in the explant medium after day 3.
4. The latent type collagenase was activated by trypsin, 4-aminophenylmercuric acetate (4-APMA), thiocyanate and deoxycholate (DOC). DOC showed irreversible dissociation of latent type enzyme in similar fashion to that exerted by 4-APMA.
5. The purified collagenase was inhibited by bovine serum, EDTA, o-phenanthroline, cysteine and dithiothreitol.抄録全体を表示PDF形式でダウンロード (1442K) -
Fumiaki SUZUKI, Yukio NAKAMURA, Yukio NAGATA, Tamiko OHSAWA, Kazuo MUR ...1981 年 89 巻 4 号 p. 1107-1112
発行日: 1981/04/01
公開日: 2008/11/18
ジャーナル フリーA pressor enzyme, renin, was purified about 60-fold by chromatography on an affinity column including pepstatin-aminohexyl-agarose with a high yield of 83 from the homogenate of adult mouse submaxillary glands. The renin obtained by the one-step purification was electrophoretically homogenous on SDS-polyacryl-amide gel and was as active as an absolutely pure renin. The renin purified by the affinity column could be separated into five active components by chromatography on CM-cellulose. Each of these renins gave a symmetrical elution profile on the CM-cellulose column and a discrete protein band on polyacrylamide gel electrophoresis at pH 8.6. Administration of nanogram quantities of each of the two major renin fractions to nephrectomized rats caused a sustained rise of blood pressure and decrease in sensitivity of the animal to angiotensin 11. This rapid and large-scale purification method using pepstatin-aminohexyl-agarose eliminates all four fractionation steps reported previously for the isolation of mouse submaxillary gland renin.抄録全体を表示PDF形式でダウンロード (1262K) -
III. Peptide Structures of the Linkage Region in Proteoglycans of Human, Porcine and Shark CartilagesMamoru ISEMURA, Tadamasa HANYU, Hiroko KOSAKA, Teruo ONO, Tokuji IKENA ...1981 年 89 巻 4 号 p. 1113-1119
発行日: 1981/04/01
公開日: 2008/11/18
ジャーナル フリーChondroitin sulfate peptidoglycans were prepared by the pronase digestion of cartilages from human knee joints, porcine tracheae, and shark crania. The β-elimination and sulfite addition reaction of these peptidoglycans yielded cysteic acid-containing peptides, and the amino acid sequences of some of them were determined. The finding that human and porcine peptidoglycans have the same sequence in the linkage region as that of the bovine peptidoglycan reported previously suggested that there is a common structural feature in the core proteins of these mammalian cartilage proteoglycans. The results also support the view that -Ser-Gly- is the minimum requisite for the formation of the xylosylserine linkage of proteochon-droitin sulfates.抄録全体を表示PDF形式でダウンロード (1515K) -
Kiyoshi NAGATA, Nobuo YOSHIDA1981 年 89 巻 4 号 p. 1121-1127
発行日: 1981/04/01
公開日: 2008/11/18
ジャーナル フリーFor simple, rapid purification of Streptomyces erythreus trypsin-like enzyme (TLE), we examined affinity chromatography with quail ovomucoid (QO) as a ligand of an affinity matrix. We prepared two affinity matrices by the cyanogen bromide method and the oxirane coupling method, and compared their binding efficiencies for TLE using frontal affinity chromatography. The affinity matrix in which QO was immobilized by the cyanogen bromide method showed 30% binding efficiency. However, another matrix, in which QO was immobilized by the oxirane coupling method with the lysine residue of the reactive site blocked by citraconylation, showed 92 binding efficiency. The results suggest that some biologically inactive QO was formed during the coupling reaction by the cyanogen bromide method. When we purified TLE from S. erythreus culture broth by affinity chromatography on QO-Sepharose, TLE was purified about 1, 100-fold by affinity chromatography, and was further purified by DEAF-Sephadex A-25 column chromatography to homogeneity. The overall yield of TLE activity was higher than 90%. Thus, we were able to greatly improve previous purification procedures.抄録全体を表示PDF形式でダウンロード (478K) -
Toshinobu FUJIYOSHI, Motoaki ANAI1981 年 89 巻 4 号 p. 1129-1136
発行日: 1981/04/01
公開日: 2008/11/18
ジャーナル フリーThe ATP-dependent deoxyribonuclease from Bacillus laterosporus has been purified to near homogeneity by a procedure involving ammonium sulfate fractionation, DEAF-cellulose chromatography, Sephadex G-150 gel filtration, DEAE-Sephadex A-25 chromatography and DNA-cellulose affinity chromatography. The purified enzyme has a molecular weight of 210, 000±8, 000 as determined by sucrose gradient sedimentation. It is composed of two nonidentical polypeptide chains with close molecular weights of around 110, 000. The substrate preference of the pure enzyme is essentially identical with the previous result obtained with the partially purified enzyme preparation (Anai, M., Mihara, T., Yamanaka, M., Shibata, T., & Takagi, Y. (1975) J. Biochem. 78, 105-114). Thus, the enzyme degrades double-stranded DNA about 100 times faster than heat-denatured DNA in the presence of ATP. Double-stranded DNA is not degraded to any measurable extent in the absence of ATP, but the enzyme exhibits activity toward denatured DNA in the absence of ATP. Furthermore, no endonuclease activity is observed on covalently closed circular duplex DNA and open circular duplex DNA.抄録全体を表示PDF形式でダウンロード (1005K) -
Toshinobu FUJIYOSHI, Juichiro NAKAYAMA, Motoaki ANAI1981 年 89 巻 4 号 p. 1137-1142
発行日: 1981/04/01
公開日: 2008/11/18
ジャーナル フリーBacillus laterosporus ATP-dependent deoxyribonuclease has been found to be inhibited by pyridoxal 5'-phosphate. The inhibition is specific for pyridoxal 5'-phosphate and pyridoxal which are required in relatively high concentrations. Pyridoxamine 5'-phosphate, pyridoxamine, and pyridoxine are ineffective. The inhibition is reversed by dilution or dialysis but can be changed to an irreversible inactivation by reduction of the enzyme. pyridoxal 5'-phosphate complex with sodium borohydride. The compound is a competitive inhibitor with respect to DNA but not ATP. Moreover, the presence of DNA substrate protects the enzyme against this inactivation but the presence of ATP shows no effect. The reduced enzyme pyridoxal 5'-phosphate complex displays a new absorption maximum at 325 nm and a fluorescence emission at 390-400 nm when excited at 325 nm which are characteristic for ε-N-(phosphopyridoxyl)lysine. Thus, B. laterosporus DNase appears to have an essential lysine residue at the DNA binding site of the enzyme, and the enzyme possesses two different active sites, a DNA binding site and an ATP binding site.抄録全体を表示PDF形式でダウンロード (410K) -
Jun NAGAI, Minoru TANAKA, Hiroshige HIBASAMI, Tadao IKEDA1981 年 89 巻 4 号 p. 1143-1148
発行日: 1981/04/01
公開日: 2008/11/18
ジャーナル フリーMepacrine at 50 μM completely protected vitamin E-deficient rat erythrocytes from peroxidative hemolysis induced by dialuric acid or reduced glutathione under the standard experimental conditions. Malondialdehyde formation, which precedes the hemolysis, was also inhibited by mepacrine. These effects of mepacrine were observed when it was added after incubating the cells with dialuric acid before the malondialdehyde formation reached 50% of its maximal value. Mepacrine also inhibited NADPH-dependent lipid peroxidation in rat liver microsomes. The degree of inhibition by mepacrine of lipid peroxidation and hemolysis was dependent on the amount of red blood cells or microsomes in the reaction mixture.抄録全体を表示PDF形式でダウンロード (419K) -
Fumio AMANO, Den'ichi MIZUNO1981 年 89 巻 4 号 p. 1149-1154
発行日: 1981/04/01
公開日: 2008/11/18
ジャーナル フリーAgglutination of phagocytic vesicles by plant lectins and lysosomes from guinea pig polymorphonuclear leukocytes (PMNs) was examined in vitro. PMNs were allowed to phagocytize paraffin oil emulsion, and phagocytic vesicles were isolated from the cells. First, the vesicles were suspended in isotonic sucrose or lactose with or without lectins, and then they were incubated at 0°C for 20min and photographed under a phase-contrast microscope. In sucrose, lectins such as Ricinus communis agglutinin, wheat germ agglutinin, Phaseolus vulgalis agglutinin-P and Ulex europeus agglutinin agglutinated the vesicles, while the vesicles without lectins remained dispersed. Concanavalin A agglutinated the vesicles in lactose solution, but the other lectins did not. These results suggest that the phagocytic vesicles have lectin receptors (carbohydrate moieties) on their cytoplasmic side, as galactosyl-, N-acetyl-galactosaminyl-, mannosyl-, glucosyl-, N-acetylglucosaminyl- and di-N-acetylchito-biose residues. Second, the phagocytic vesicles were incubated with lysosomes. Lysosomes induced agglutination of the vesicles immediately after incubation and this agglutination was inhibited by simultaneous addition of 50mM mannose, fucose, N-acetylglucosamine, lactose and maltose, and 7mM N-acetylneuraminic acid and 1.25mg/ml fetuin. The results show that lysosomes agglutinated the phagocytic vesicles in vitro, and suggest that this interaction is mediated by recognition of the carbohydrate moieties on the vesicles.抄録全体を表示PDF形式でダウンロード (1879K) -
Takashi NAGASAWA, Ryuhei FUNABIKI1981 年 89 巻 4 号 p. 1155-1161
発行日: 1981/04/01
公開日: 2008/11/18
ジャーナル フリーNτ-Methylhistidine (3-methylhistidine) in urine of the rat is mainly derived from the degradation of actin and myosin in skeletal muscle, intestine and skin. The fractional degradation rates of the myosin-actin pools of these tissues were calculated from the time course of increase in the specific radioactivities of Nτ-methylhistidine after daily administration of [methyl-14C] methionine to young adult rats under conditions of restricted food intake. The contributions to urinary excretion of Nτ-methylhistidine from the three tissues were calculated from the fractional degradation rates and Nτ-methylhistidine contents of the three tissues; 75.6% for skeletal muscle, 2.2% for intestine and 22.2% for skin. The results show that the skeletal muscle is the major source of urinary Nτ-methylhistidine output, but the contribution of skin is not negligible in rats. The specific radioactivity of Nτ-methylhistidine in urine was much higher than that of skeletal muscle. The fractional degradation rates of myosin and actin in skeletal muscle had similar values. Although the specific radioactivities of Nτ-methylhistidine in myosin and actin were very different, the mean value was similar to that in mixed skeletal muscle.抄録全体を表示PDF形式でダウンロード (507K) -
Keizo TESHIMA, Kiyoshi IKEDA, Kozo HAMAGUCHI, Kyozo HAYASHI1981 年 89 巻 4 号 p. 1163-1174
発行日: 1981/04/01
公開日: 2008/11/18
ジャーナル フリーThree cobra phospholipases A2 of N. naja siamensis, N. naja kaouthia, and N. naja ctra showed similar monomer-dimer equilibria at neutral pH values. The CD spectra in the aromatic and far-ultraviolet regions were almost the same for the three enzymes. Bindings of n-alkylphosphorylcholines were studied by employing the CD spectral changes of the enzymes under conditions where they exist in the monomeric form. On adding the substrates at concentrations below the critical micelle concentration (cmc), positive ellipticities in the aromatic region significantly decreased, but abruptly increased when the concentrations were increased above the cmc. The negative ellipticities in the far-ultraviolet region, reflecting the backbone conformations of proteins, increased in the presence of the substrates below the cmc but did not significantly change above the cmc. The binding constants determined from the changes in the aromatic and far-ultraviolet regions were in good agreement with each other. All three cobra enzymes gave almost the same binding constants.
Over a wide pH range of 2.5 to 8.5, the binding constants of the substrates were almost independent of pH, and were hardly affected by Ca2+ ion binding, although His 48 and Asp 49 with pK values of 7.55 and 5.4, respectively, in the active site are perturbed by Ca2+ ion binding (Teshima et al. (1981) J. Biochem. 89, 13-20).
The chain-length dependence of the binding constant of n-alkylphosphoryl-choline was studied. The contribution of one methylene moiety to the unitary free energy of the binding was determined to be about 0.39 kcal/mol at 25°C, which is larger than that, 0.26 kcal/mol, for the enzyme of A. halys blomhoffii (Ikeda & Samejima, submitted to J. Biochem.), and slightly smaller than that for porcine pancreas enzyme, about 0.5 kcal/mol, calculated from the data of van Dam-Mieras et al. ((1975) Biochemistry 14, 5387-5394) and of Verheij et al. ((1980) Biochemistry 19, 743-750). This suggests that the substrate binding sites of cobra phospholipases A2 are as hydrophobic as that of the porcine enzyme, but are more hydrophobic than that of the A. halys blomhoffii enzyme.抄録全体を表示PDF形式でダウンロード (759K) -
Kiyoshi IKEDA, Yuji SAMEJIMA1981 年 89 巻 4 号 p. 1175-1184
発行日: 1981/04/01
公開日: 2008/11/18
ジャーナル フリーBindings of monodispersed n-dodecyl-, n-decyl-, and n-octylphosphorylcholines (n-C12PC, n-C10PC, and n-C8PC, respectively) to phospholipase A2-II of A. halys blomhoffii were studied by the aromatic circular dichroism (CD) or ultraviolet difference absorption method. The binding constants to the apoenzyme at around 25°C, pH 6.4-6.5, and ionic strength 0.1 were 1600-1800, 820, and 320M-1 for n-C12PC, n-C18PC, and n-C8PC, respectively. The contribution of one methylene moiety in the alkyl chain of the substrate to the unitary free energy of binding was calculated to be 0.26 kcal/mol, which is smaller than that for porcine pancreatic enzyme, about 0.5 kcal/mol (van Dam-Mieras et al. (1975) Biochemistry 14, 5387-5394 and Verheij et al. (1980) Biochemistry 19, 743-750) and that for cobra enzymes, 0.39 kcal/mol (Teshima et al., submitted to J. Biochem.). This suggests that the substrate binding site of phospholipase A2-II of A. halys blomhoffii is less hydrophobic than those of the enzymes of porcine pancreas and cobra venoms.
Binding of Ca2+ to phospholipase A2-II was studied by the aromatic circular dichroism method. The binding constants at pH 6.53 and 8.32 were 1880 and 6300M-1, respectively. If we assume that the pK value of His 48 in the active site is around 7, the result indicates a pK shift of His 48 by about 0.5 pH unit to the acidic side on binding of Ca2+.
The binding constants of n-C12PC to the apoenzyme and Ca2+ complex at pH 8.2-8.4 were smaller than those at pH 6.4-6.5 by a factor of about two. This suggests a pK shift of His 48 by about 0.3 pH unit to the alkaline side on binding of the substrate.
The binding constant of n-C12PC to the Ca2+-enzyme complex was smaller than that for the apoenzyme by a factor of about 2.5 to 2.7 at pH 8.2-8.4 and 6.4-6.5. This indicates that an electrostatic interaction between the bound Ca2+ ion and the negatively charged phosphate moiety of the substrate is not important in this case.抄録全体を表示PDF形式でダウンロード (571K) -
Fuyuhiko INAGAKI, Yoshio KAWANO, Ichio SHIMADA, Kenji TAKAHASHI, Tatsu ...1981 年 89 巻 4 号 p. 1185-1195
発行日: 1981/04/01
公開日: 2008/11/18
ジャーナル フリーThe 270-MHz 1H NMR spectra and fluorescence of ribonuclease T1 and carboxy-methylated ribonuclease T1 were measured in aqueous solution. Histidine C4 proton resonances were assigned to individual residues. From the pH dependences of the chemical shifts of histidine C2 and C4 protons, the pKa, values of histidine residues were obtained by the non-linear least-squares method. The hydrogen→deuterium exchange rates of histidine C2 protons were determined as a measure of the accessibility of histidine residues to the solvent. Each histidine residue of ribonuclease T1 was found to interact with a carboxylate group of an aspartic or glutamic acid residue; in particular, His-40 was shown to interact with Glu-58. Upon carboxy-methylation of Glu-58, His-92 and His-27 are more shielded from the solvent while His-40 remains exposed to the solvent. The 67.9-MHz 13C NMR spectra were measured for the 13C-enriched preparation of carboxymethylated ribonuclease T1. From the pH dependence of 13C chemical shift, the pKa value of the carboxymethylated Glu-58 was found to be unusually low, suggesting the formation of an ionic or hydrogen bond between this carboxymethyl group and a positively charged group, possibly of Arg-77.抄録全体を表示PDF形式でダウンロード (749K) -
Eisuke NISHIDA1981 年 89 巻 4 号 p. 1197-1203
発行日: 1981/04/01
公開日: 2008/11/18
ジャーナル フリーA 94, 000-dalton protein that has been shown to modulate microtubule assembly in a Mg2+- or Ca2+-dependent manner (Nishida & Sakai (1980) J. Biochem. 88, 1577-1586) was here shown to inhibit actin polymerization. The protein factor inhibited the rate and the extent of actin polymerization under nearly physiological conditions (for example, in 3mM MgCl2 plus 90mM KCl at pH 6.8). The inhibitory effect was dependent on divalent cation concentration; the lower the Mg2+ concentration was, the weaker the inhibitory effect. The inhibition was stoichiometric; addition of the protein factor caused a linear decrease in the extent of actin polymerization as measured by the viscosity increase, and under optimal conditions for inhibition about an equimolar amount of the protein factor was sufficient to inhibit the actin polymerization completely. Furthermore, inclusion of the protein factor increased the critical concentration of actin required for polymerization by a concentration nearly equivalent to that of the added factor. These results suggest the formation of a 1:1 complex between actin and the protein factor, which does not polymerize at all. Another assay for actin polymerization, pelleting of actin filaments by ultra-centrifugation, confirmed the inhibitory effect of the protein factor. In addition to the inhibitory effect on polymerization, the protein factor had the ability to depoly-merize actin filaments. We have temporarily called this protein factor PI factor. It may play an important role in cell structure and function through its interactions with actin and microtubules.抄録全体を表示PDF形式でダウンロード (511K) -
Takayoshi WAKAGI, Takahisa OHTA1981 年 89 巻 4 号 p. 1205-1213
発行日: 1981/04/01
公開日: 2008/11/18
ジャーナル フリーThe results of studies on the initial velocity in hydrolysis reactions of ATP and other nucleoside triphosphates with beef liver mitochondrial ATPase can be summarized as follows.
1. Double reciprocal plots of substrate concentration vs. initial velocity were linear for all the nucleoside triphosphates tested except for ATP, which showed a negative cooperativity.
2. Bicarbonate ion increased the rate of ATP hydrolysis and diminished its negative cooperativity, whereas hydrolysis of other nucleoside triphosphates was only slightly affected by the anion.
3. An excess of nucleoside triphosphate apparently inhibited its own hydrolysis for all kinds of nucleoside triphosphates tested, whereas an excess of magnesium ion apparently inhibited only ATP hydrolysis.
4. Inhibition of ATPase activity with an excess of magnesium ion was no longer observed when the reaction was carried out at low temperature (10°C) or in the presence of sulfate. Under these conditions, the kinetics of ATP hydrolysis were apparently of simple Michaelis-Menten type.
These observations suggest the existence of two states (“A” and “N”) of beef liver mitochondrial ATPase. The state “A” is characterized by phenomena specifically affecting ATP hydrolysis, such as the inhibition by excess magnesium ion, and the negatively cooperative profile in the dose-response curve of ATP. In the state “N” proceed the hydrolysis reactions of other nucleoside triphosphates and of ATP under limited conditions. The two states (“A” and “N”) can be related to an enzyme model with a catalytic site and a regulatory site. Computor simulation revealed that such a model could account well for the experimental data.抄録全体を表示PDF形式でダウンロード (645K) -
Ewa PRÒCHNIEWICZ, Toshio YANAGIDA1981 年 89 巻 4 号 p. 1215-1221
発行日: 1981/04/01
公開日: 2008/11/18
ジャーナル フリーComparison of interactions between the monomers of chicken gizzard and skeletal muscle actin revealed: 1. A more pronounced increase of the extent of polymerization of chicken gizzard than skeletal muscle actin with temperature, which resulted in higher positive changes of entropy and enthalpy of the former than of the latter species. 2. A difference in spectral changes accompanying polymerization: the changes at 295nm, attibuted to environmental changes around tryptophan residues, were less pronounced for gizzard than for skeletal actin. 3. A difference in the amount of heavy meromyosin added to gizzard and to skeletal F-actin, with which the degree of flow birefringence of the acto-HMM complex is minimum: this amount was lower in the former than in the latter case. These results indicate quantitative differences between intermonomer interactions involved in polymerization of both actin species and also a possible difference in cooperativity between the monomers within the polymers of gizzard and skeletal actin.抄録全体を表示PDF形式でダウンロード (516K) -
Naofumi KITABATAKE, Ryuzo SASAKI, Hideo CHIBA1981 年 89 巻 4 号 p. 1223-1229
発行日: 1981/04/01
公開日: 2008/11/18
ジャーナル フリーEvidence supporting the existence of three aldehyde dehydrogenases in bovine liver has been confirmed and the immunological properties of these isozymes have been compared.
1. The cytoplasmic and mitochondrial aldehyde dehydrogenases were distinguished in size by subjecting the bovine liver crude extract to Sephadex G-150 chromatography. The molecular weight of the cytoplasmic aldehyde dehydrogenase was determined to be 230, 000 daltons, larger than the mitochondrial enzyme by approxi-mately 15, 000 daltons.
2. Submitochondrial fractionation indicated that the mitochondrial aldehyde dehydrogenase was located in the intermembrane space.
3. Anti-mitochondrial aldehyde dehydrogenase antibody from rabbit gave an immunoreaction line not only with the mitochondrial enzyme but also with the cytoplasmic enzyme, and inhibited the activity of both enzymes. The anti-cytoplasmic aldehyde dehydrogenase antibody reacted with the cytoplasmic enzyme but had no effect on the mitochondrial enzyme. Neither antibody reacted with the microsomal aldehyde dehydrogenase solubilized with sodium deoxycholate.抄録全体を表示PDF形式でダウンロード (1184K) -
Kouichi MIYATA, Masahira NAKAMURA, Katsumi TOMODA1981 年 89 巻 4 号 p. 1231-1237
発行日: 1981/04/01
公開日: 2008/11/18
ジャーナル フリーSerratia protease (TSP) [EC 3. 4. 24] was bound stoichiometrically to α2macroglobulin (α2M), which was purified and crystallized from human plasma, but apo TSP was not bound. On formation of the TSP-α2M complex the enzymatic activity of the bound TSP was affected with respect to substrates; Km values of the bound TSP were unchanged but Vmax values were reduced. α2M was cleft at the mid-region of its subunits chains by TSP, which resulted in a conformational change of the α2M molecule with TSP.抄録全体を表示PDF形式でダウンロード (418K) -
I. Effect of an Artificially Imposed H+ Gradient on Ca2+ UptakeTakashi UENO, Takamitsu SEKINE1981 年 89 巻 4 号 p. 1239-1246
発行日: 1981/04/01
公開日: 2008/11/18
ジャーナル フリーTo ascertain the function of H+ flux in active Ca2+ transport into sarcoplasmic reticulum vesicles, the effect of pH gradient on Ca2+ transport was examined.
A transient H+ gradient (inside-acidic) was imposed on K+-loaded sarcoplasmic reticulum vesicles with the aid of K+-H+ exchange driven by nigericin. This proton gradient was dessipated rapidly and concomitantly with ATP-driven Ca2+ transport. Under these conditions, the initial rate of the Ca2+ uptake was increased about 1.5-fold. The stimulation of Ca2+ uptake was completely lost when the pH gradient was cancelled with an uncoupler plus membrane permeable cation before Ca2+ uptake.
These results are interpreted in terms of H+ efflux coupled with Ca2+ transport.抄録全体を表示PDF形式でダウンロード (543K) -
II. H+ Ejection during Ca2+ UptakeTakashi UENO, Takamitsu SEKINE1981 年 89 巻 4 号 p. 1247-1252
発行日: 1981/04/01
公開日: 2008/11/18
ジャーナル フリーProton efflux during Ca2+ transport into sarcoplasmic reticulum vesicles was examined.
Although a rapid H+ ejection was observed during the initial phase of Ca2+ uptake and the amount of the liberated H+ was more than that due to hydrolysis of ATP, generation of a pH difference as a result of the H+ efflux could not be detected by direct pH measurement with a pH meter.
Alkalinization of the inside of the vesicles during Ca2+ uptake was more precisely examined by flow dialysis assay and a significant uptake of acetate or salicylate into the vesicles was found, suggesting the generation of a small pH difference across the SR membrane.
From these results, it was concluded that counter-transport of H+ was operative in Ca2+ uptake but that only a relatively small pH difference was generated as a result of the H+ efflux.
The intrinsic buffering capacity of sarcoplasmic reticulum vesicles was measured and a relatively large value (130 nmol H+/pH unit/mg at pH 6.2) was obtained.抄録全体を表示PDF形式でダウンロード (459K) -
Masao IWAMORI, Yoshitaka NAGAI1981 年 89 巻 4 号 p. 1253-1264
発行日: 1981/04/01
公開日: 2008/11/18
ジャーナル フリーRabbit skeletal muscle contained 28.4 nanomol/g wet weight of lipid-bound sialic acid, and 73.4% of the total lipid-bound sialic acid was recovered in a monosialo-ganglioside fraction. Monosialoganglioside components were isolated and the structures were determined by exoglycosidase treatment and permethylation analysis. The major monosialoganglioside was GM3 (73.2% of monosialogangliosides), and gangliosides of the lacto-series with at least three repeating units of lactosamine, Gal (β, 1-4)G1cNAc(β, 1-3), were identified. A newly found ganglioside was N-acetylneuraminosyl lacto-N-noroctaosyl ceramide, NeuAc(α, 2-3) Gal (β, 1-4) GlcNAc (β, 1-3) Gal (β, 1-4) GlcNAc (β, 1-3) Gal (β, 1-4) GlcNAc (β, 1-3) Gal (β, 1-4) Glc (β, 1-1) ceramide. The amounts of lacto-series gangliosides in the monosialoganglioside fraction were as follows: sialosyl lacto-N-neotetraosyl ceramide (9.8%), sialosyl lacto-N-norhexaosyl ceramide (11.9%) and sialosyl lacto-N-noroctaosyl ceramide (5.1%). N-Glycolylneuraminic acid was found in sialosyl lacto-N-neotetraosyl ceramide and sialosyl lacto-N-norhexaosyl ceramide but the amount was less than 0.2%. The mobilities of lacto-series gangliosides were compared with those of ganglio-series gangliosides. Fatty acid and long chain base compositions were also determined.抄録全体を表示PDF形式でダウンロード (2889K) -
Hiroshi TAKADA, Yoshio ARAKI, Eiji ITO1981 年 89 巻 4 号 p. 1265-1274
発行日: 1981/04/01
公開日: 2008/11/18
ジャーナル フリーThe structure of polygalactosamine purified from the culture fluid of Aspergillus parasiticus AHU 7165 was studied. Partial acid hydrolysis of this polysaccharide, in which 55 to 65% of the monosaccharide residues were N-unsubstituted, gave a series of galactosamine oligosaccharides (dimer to hexamer) in a good yield. From the data on analyses of the polysaccharide and its oligosaccharides by gel filtration, periodate oxidation, methylation, and proton NMR measurement, the polysac-charide was characterized as a linear chain of α(1-4)-linked galactosamine residues. The N-unsubstituted galactosamine residues are probably distributed in a random fashion over the polysaccharide chain.抄録全体を表示PDF形式でダウンロード (776K) -
Isao UNO, Tatsuo ISHIKAWA1981 年 89 巻 4 号 p. 1275-1281
発行日: 1981/04/01
公開日: 2008/11/18
ジャーナル フリーA single cAMP-receptor protein could be detected in mycelial extracts of Copriuus macrorhizus by using the photoaffinity cAMP-analogue, 8-N3-cAMP. The protein which specifically bound 32P-labeled 8-N3-cAMP had an apparent molecular weight of 46, 000 as determined by an SDS-polyacrylamide gel elctrophoresis system. The 46, 000-dalton protein was characterized by the dissociation constant for [32P]-8-N3-cAMP, and by the nucleotide specific inhibition of [32P]-8-N3-cAMP binding. The 46, 000-dalton protein was co-chromatographed on a DEAE-cellulose column with cAMP-dependent protein kinase. The levels of [32P]-8-N3-cAMP-binding and protein kinase activities in mycelial extracts of strains used was always in parallel. The result indicated that the 46, 000-dalton protein may be a regulatory subunit of protein kinase with the capacity to bind cAMP. cAMP-dependent protein kinase of this fungus was immunologically different from those of higher animals.抄録全体を表示PDF形式でダウンロード (1148K) -
Ken-Ichi FURUKAWA, Akio INOUE, Yuji TONOMURA1981 年 89 巻 4 号 p. 1283-1292
発行日: 1981/04/01
公開日: 2008/11/18
ジャーナル フリーWe [Kanazawa, T. & Tonomura, Y. (1964) J. Biochem. 57, 604-615; Furukawa, K. -I., Ikebe, M., Inoue, A., & Tonomura, Y. (1980) J. Biochem. 88, 1629-1641] previously reported that the initial Pi-burst size of the myosin-ATPase reaction was about 11 mol/mol Myo head at 1 μm of free Mg2+ ions which decreased with an increase in the free Mg2+ concentration, but was still larger than the stoichiometric value (0.5 mol/mol Myo head) at 10mM of Mg2+ ions. We attributed the Pi-burst size larger than the stoichiometric value to a so-called extra Pi burst. The results of our present study support our hypothesis that the larger burst size is due to contamination of the true Pi burst by an extra Pi burst.
The rate of fluorescence enhancement of myosin induced by ATP increased with an increase in the free Mg2+ concentration, while the final extent of fluorescence enhancement in the steady state was independent of the free Mg2+ concentration (1 μM to 10mm). When excess EDTA was added to the reaction mixture in the steady state, the fluorescence decreased to the original level. The rate of decay was independent of the free Mg2+ concentration which had been used for the fluorescence enhancement. Furthermore, the time course of fluorescence decay was equal to that of the transition of ATPase from the Mg2+ to the EDTA(K+) type.
The amount of free Pi liberated (free Pi) was measured by a double-membrane filtration method at various free Mg2+ concentrations, and compared with the amount of Pi liberated when the reaction was stopped by addition of TCA (TCA-Pi). The steady-state rates of free Pi liberation were equal to those of TCA-Pi liberation at various free Mg2+ concentrations. The difference between the amounts of TCA-Pi and free Pi, i.e., the amount of P bound to myosin, was 0.49-0.51 mol/ mol Myo head regardless of the free Mg2+ concentration. Furthermore, at the completion of the initial Pi burst, the amount of ADP bound to myosin was 0.48-0.50 mol/mol Myo head at various free Mg2+ concentrations.抄録全体を表示PDF形式でダウンロード (697K) -
Naoya KENMOCHI, Kunio TSURUGI, Kikuo OGATA1981 年 89 巻 4 号 p. 1293-1308
発行日: 1981/04/01
公開日: 2008/11/18
ジャーナル フリーBasic proteins of 40S and 60S subunits of Artemia salina and rat liver were analyzed using two different systems of two-dimensional acrylamide gel electrophoresis. In system I, the first dimension was run at pH 8.6 and the 2nd at pH 4.9, and in system II, the first was run at pH 8.6 and the 2nd in the presence of SDS, “Three-dimensional” electrophoresis was further used for the identification of individual ribosomal proteins.
1. Artemia salina 40S proteins were separated into 27 proteins by “three-dimensional” gel electrophoresis. Their two-dimensional electrophoretogram in system I was somewhat different from that of liver 40S subunits, especially in the less basic region near the origin. Individual Artemia salina 40S proteins were designated according to their correspondence to 40S proteins of rat liver. The proteins S8 or S9 of both subunits overlapped between the two species. Proteins corresponding to rat liver S3b, S5a, and S5 proteins were not detected in Artemia salina 40S proteins. Forty-S proteins of Artemia salina and rat liver were further analyzed by two-dimensional gel electrophoresis in system IT. S7, S9, and S29 proteins overlapped between the two species. Thus, S9 protein may have been almost completely conserved during evolution.
2. Thirty-eight proteins were identified in Artemia salina 60S proteins by “three-dimensional” electrophoresis. The pattern was different from that of rat liver, especially in the basic region on two-dimensional gels in systems I and II. Although the correspondence of individual proteins between the two species was very difficult to find, we designated Artemia salina 60S proteins by considering the mutual relationship to rat liver 60S proteins on the gel. L5, L31, L18, and L18a proteins of Artemia salina appeared to shift to more acidic regions as compared with corresponding 60S proteins of rat liver.
3. The number averages of molecular weights of 40S proteins were 18, 600 for Artemia salina (27 proteins) and 18, 500 for rat liver (29 proteins). Those of 60S proteins were 21, 600 for Artemia salina (38 proteins) 21, 800 for rat liver (38 proteins). The averages of relative basicities of 40S proteins were similar in both species, whereas the average of those of 60S proteins of Artemia salina was lower than that of rat liver 60S proteins.
4. The protein moiety of 40S ribosomes has been conserved more than that of 60S ribosomes during the evolution of animal species.抄録全体を表示PDF形式でダウンロード (3951K) -
Masahisa NAKAMURA, Akiya HINO, Ikuo YASUMASU, Junzo KATO1981 年 89 巻 4 号 p. 1309-1315
発行日: 1981/04/01
公開日: 2008/11/18
ジャーナル フリーLactate markedly increased the rate of [3H]leucine incorporation into the protein of isolated round spermatids (steps 1-8) from rat testes. Four kinds of hexoses, glucose, fructose, galactose, and monnose, also stimulated [3H]leucine incorporation, but to much lesser extents than lactate. Ribose had no effect. The glucose-induced stimulation of protein synthesis was entirely suppressed by iodoacetate and NaF, whereas iodoacetate and NaF were without effect on the lactate-induced increase in protein synthesis. Lactate stimulated both protein synthesis and ATP production in the spermatids. However, both of these stimulatory effects of lactate were completely blocked by DNP and rotenone. Rotenone entirely blocked oxygen consumption, as expected, whilst DNP enhanced it additively with lactate. Moreover, lactate was without influence on either transport of α-[3H]AIB into spermatids or incorporation of [3H]leucine into protein of a cell-free system of spermatids.
These findings suggest that lactate may increase the protein synthesis of spermatids in the same fashion as glucose, and that the effect of lactate in increasing the level of ATP during incubation in vitro may be a major factor in the mechanism of stimulation of protein synthesis in the spermatids.抄録全体を表示PDF形式でダウンロード (507K) -
Osamu ODA, Takashi MANABE, Tsuneo OKUYAMA1981 年 89 巻 4 号 p. 1317-1323
発行日: 1981/04/01
公開日: 2008/11/18
ジャーナル フリーWe examined whether bovine serum amine oxidase (BSAO) was able to oxidize lysyl peptides. Seven synthetic peptides, Z-Lys-Leu-OMe, Z-His-Lys-Leu-OMe, Z-Val-Leu-Gly-Lys-Leu-OMe, Ala-Ala-Lys, Ala-Lys-Ala, Phe-Lys, and Gly-Lys were incubated with BSAO at 37°C for 4 days, and the extent of oxidation (H2O2 formation) was assayed by o-dianisidine method. The amino acid sequence of lysyl peptides affected the oxidation rate by BSAO. The rate of oxidation of Z-Lys-Leu-OMe was about fifty fold higher than that of Z-His-Lys-Leu-OMe. Z-Lys-Leu-OMe, the lysyl peptide most reactive with BSAO was analyzed by the 3-methyl-2-benzothiazolinone hydrochloride method, amino acid analysis, and the 2, 4, 6, -trinitrobenzene sulfonic acid method. The production of aldehyde, a decrease in the lysine/leucine ratio, and a decrease in ε-amino group were confirmed. These results show that bovine serum amine oxidase is able to oxidize the s-amino group of lysyl peptides.抄録全体を表示PDF形式でダウンロード (463K) -
Tatsuya SAMEJIMA, Tsuneo MIYAHARA, Atsushi TAKEDA, Akira HACHIMORI, Ko ...1981 年 89 巻 4 号 p. 1325-1332
発行日: 1981/04/01
公開日: 2008/11/18
ジャーナル フリーPorcine erythrocyte catalase [EC 1. 11. 1. 6] (molecular weight, ca. 250, 000) was found to dissociate partially between pH 3.5 and 3.0 and completely below pH 3.0 into two presumably identical 1/2-sized subunits (molecular weight, 112, 000) as estimated by ultracentrifugal analyses. This dissociation was accompanied by a marked change in hydrodynamic properties; the sedimentation coefficient decreased from about 11S to 4S. This acid denaturation also resulted in complete loss of enzyme activity and disappearance of absorption bands characteristic of heme protein, in particular, a shift of the Soret band from 405nm to a small and broad band at 375 nm. The change in enzyme activity correlated well with that of the Soret band, depending on the denaturation time and pH used. Reversible recovery of enzyme activity was not detected below pH 3.1 after 2h denaturation. The pH dependence of α-helical content estimated from the CD intensity at 222nm also correlated well with that of enzyme activity. The rate constants of initial reaction of acid denaturation at several pHs were determined by following the changes in the Soret band with time, since the changes showed an isosbestic point at 384nm. The results revealed that the first-order rate constant at pH 3.0 was 45 times larger than that at pH 3.4, indicating that the rate of acid denaturation increased rapidly within a narrow acidic pH range. The temperature dependence of the denaturation rate was also measured and the activation energy for the acid denaturation was found to be 76.3 kcal/mol from an Arrhenius plot.抄録全体を表示PDF形式でダウンロード (642K) -
Takao OJIMA, Kiyoyoshi NISHITA, Shizuo WATANABE1981 年 89 巻 4 号 p. 1333-1335
発行日: 1981/04/01
公開日: 2008/11/18
ジャーナル フリーMyosin from striated adductor muscle of “Akazara” scallop was incubated at 30°C for 5min in a medium containing 2mM MgCl2 and various concentrations of Ca2+ ions. It was observed that the 30°C-treatment resulted in a decrease in the Ca2+-sensitivity of myosin-ATPase as well as in the release of the regulatory light chain (EDTA-LC) of myosin. The 30°C-treated myosin was then subjected to a cooling treatment, being kept for 18 h at 0°C. It was found that EDTA-LC recombined with myosin and that Ca2+-sensitivity of myosin-ATPase was restored.
It was also found that Ca2+ alone was about 70 times more effective than Mg2+ alone in preventing the heat-induced release of EDTA-LC from occurring and also in recombination of EDTA-LC with the heat-treated myosin.抄録全体を表示PDF形式でダウンロード (819K) -
Koscak MARUYAMA, Hiroshi SAKAI1981 年 89 巻 4 号 p. 1337-1340
発行日: 1981/04/01
公開日: 2008/11/18
ジャーナル フリーA motility-regulatory protein called cell β-actinin was isolated from rat kidney cytosol. Although its molecular weight (_??_80, 000) is different from that of β-actinin from skeletal muscle (heterodimer of 34, 000 and 37, 000), the action on actin is the same for both. Cell β-actinin accelerates polymerization of actin by enhanced nucleation resulting in shorter actin filaments, inhibits the reassociation of sonicated F-actin fragments, and forms a Mg polymer in the presence of Mg ions. Cell β-actinin appears to be present in various tissues of rat, rabbit, chicken, and pig. Cell β-actinin is thought to function as an accelerator of actin polymerization in nonmuscle cells, in which actin monomers are abundantly present.抄録全体を表示PDF形式でダウンロード (1561K) -
Issei MABUCHI1981 年 89 巻 4 号 p. 1341-1344
発行日: 1981/04/01
公開日: 2008/11/18
ジャーナル フリーA protein which is capable of depolymerizing F-actin was purified from an extract of unfertilized starfish eggs by the use of DEAE-cellulose column chromatography and hydroxylapatite column chromatography. This protein has an apparent molecular weight of 17, 000. It inhibited the extent of actin polymerization as well as depolymerizing F-actin rapidly. It was shown that this protein reacts with actin at a molar ratio of 1:1. The properties of this protein were compared with those of profilin from mammalian tissues.抄録全体を表示PDF形式でダウンロード (950K) -
1981 年 89 巻 4 号 p. 1345
発行日: 1981年
公開日: 2008/11/18
ジャーナル フリーPDF形式でダウンロード (21K)
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