巻号一覧
117 巻, 5 号
選択された号の論文の36件中1～36を表示しています
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Takashi Kumasaka, Hiroyuki Takeya, Masaki Yamamoto, Yoshio Yamakawa, T ...1995 年 117 巻 5 号 p. 929-930
発行日: 1995年
公開日: 2008/11/18
ジャーナル フリーH2-proteinase, a non-hemorrhagic metalloproteinase from the venom of Trimeresurus flavoviridis, has been crystallized by vapor diffusion from solutions containing ammonium sulfate. The crystals belong to the tetragonal space group, P41212 or P43212, with unit cell dimensions of a=b=77.8 Å and c=82.3 Å. The asymmetric unit contains one protein molecule. Diffraction data for a native crystal were collected up to 2.0 Å resolution.抄録全体を表示PDF形式でダウンロード (1411K) -
Chikahiro Nagata, Noriyuki Igarashi, Hideaki Moriyama, Taketomo Fujiwa ...1995 年 117 巻 5 号 p. 931-932
発行日: 1995年
公開日: 2008/11/18
ジャーナル フリーCytochrome c-554 from Nitrosomonas europaea was crystallized by the batch method utilizing ammonium sulfate as the precipitant. The crystals belong to the space group, P21 with unit cell dimensions of a=64.8 Å, b=44.7 Å, c=78.7 Å, and β=99.0°. The crystals diffracted X-rays beyond 2.5 Å resolution.抄録全体を表示PDF形式でダウンロード (1374K) -
Masateru Nishihara, Yosuke Koga1995 年 117 巻 5 号 p. 933-935
発行日: 1995年
公開日: 2008/11/18
ジャーナル フリーOne of the most characteristic features of archaebacterial ether phospholipids is the enantiomeric configuration of their glycerophosphate backbone (sn-glycerol-1-phosphate), that is the mirror image of the structure of the eubacterial or eukaryotic counterpart. The enzyme that forms glycerophosphate of this configuration was found for the first time in a cell-free extract of the methanogen, Methanobacterium thermoautotrophicum, and was identified as sn-glycerol-1-phosphate: NAD+ oxidoreductase (sn-glycerol-1-phosphate dehydrogenase) after partial purification. Because sn-glycerol-1-phosphate has been found to be a precursor of ether lipids of this organism, sn-glycerol-1-phosphate dehydrogenase is a key enzyme in the biosynthesis of the enantiomeric ether lipids of methanogens.抄録全体を表示PDF形式でダウンロード (383K) -
Hidehiko Kikuchi, Shinji Sugiyama, Shinobu Imajoh-Ohmi1995 年 117 巻 5 号 p. 936-939
発行日: 1995年
公開日: 2008/11/18
ジャーナル フリーThe Fas antigen (CD95) is a receptor for apoptotic cell death signals. Human T Jurkat cells that express the Fas molecule on the cell surface are sensitive to cytotoxic anti-Fas monoclonal antibodies (mAbs). We report here that fetal calf serum (FCS) contains a novel factor (SAF: _??_erum _??_nti-apoptosis _??_actor) that suppresses Fas-mediated apoptosis of Jurkat cells. The apoptotic cell death of Jurkat cells induced by an anti-Fas mAb was accelerated when FCS was removed from the medium. Depletion of serum itself did not induce apoptosis without the mAb. The apoptosis-suppressing activity was not seen in neonatal calf or adult bovine serum, suggesting that the SAF activity was developmentally regulated. The anti-apoptosis activity was not detected in any fraction of FCS obtained on gel filtration chromatography. However, the activity was recovered in a certain fraction on the addition of calf serum, suggesting that the anti-apoptotic activity of SAF requires other serum factor(s). We purified SAF from FCS using an assay system reconstituted with calf serum, and raised antibodies (Abs) to it in rabbits. The anti-SAF Ab inhibited the apoptosis-suppressing activity, indicating that SAF is a key factor for the anti-apoptotic activity of FCS.抄録全体を表示PDF形式でダウンロード (1516K) -
Condensation of Collagen Fibrils to the Direct Vicinity of Fibroblasts as a Cause of Gel ContractionMasayuki Yamato, Eijiro Adachi, Kiyotaka Yamamoto, Toshihiko Hayashi1995 年 117 巻 5 号 p. 940-946
発行日: 1995年
公開日: 2008/11/18
ジャーナル フリーFibroblasts were cultured within FITC-prelabeled type I collagen. Cells initially round in shape protruded processes, and began to collect the fibrils into their vicinity. Repeated protrusion and withdrawal of cell processes was observed. Consequently, condensed fluorescence was observed on the elongated bipolar cells stained with rhodamine-phalloidin. Concomitantly with these events, the gel began to contract in overall size, with an increase of fluorescence intensity. Scanning electron micrographs of the contracted gel showed a disproportional distribution of collagen fibrils: a highly condensed region surrounding cell bodies and a moderately condensed region. A major portion of condensed fibrils may have been derived from reconstituted collagen fibrils, since fibroblasts within collagen gel synthesized little collagen. When the gel adhered to glass tightly, so that overall contraction was prevented, the fluorescence in a range of scores of micrometers from the cells disappeared owing to depletion of fibrils by the cells. The combined spaces with null fluorescence in total under repressed contraction corresponded well to the reduction in volume due to gel contraction. It seems likely that the fibril condensation onto the cells causes the overall gel contraction.抄録全体を表示PDF形式でダウンロード (5395K) -
Masayuki Takahashi, Bengt Nordén1995 年 117 巻 5 号 p. 947-951
発行日: 1995年
公開日: 2008/11/18
ジャーナル フリーThe ability to accommodate three single-strands of DNA by each filamentous RecA-DNA complex, in the presence of the non-hydrolysable ATP analog, adenosine 5'-O-3-thiotriphosphate (ATP7S), was demonstrated by monitoring the fast renaturation that occurs between the complementary homopolymers, poly (dA) and poly (dT), upon sodium dodecyl sulfate (SDS)-induced dissociation of RecA. The renaturation could be followed from the linear dichroism signal of DNA. The reaction was fast and complete when SDS was added to a mixture of RecA having three nucleobases per subunit each of poly (dA) and poly(dT), while the reaction was slower and incomplete in the absence of RecA. The efficient renaturation indicates that the two DNA strands were bound close to each other, i.e., to the same RecA filament. Similar fast renaturation was observed upon the addition of SDS to a mixture of RecA with three bases each of poly (dA), poly (dT), and a non-complementary DNA, poly (dC), even when one of the complementary DNAs was added last as the third strand. Since the dissociation of DNA from RecA is extremely slow in the presence of ATP-γS, and thus no renaturation can occur within the RecA filaments, the results indicate that RecA can bind three single-stranded DNA molecules.抄録全体を表示PDF形式でダウンロード (641K) -
Yingjie Yu, Kazutoyo Terada, Akitoshi Nagasaki, Masaki Takiguchi, Masa ...1995 年 117 巻 5 号 p. 952-957
発行日: 1995年
公開日: 2008/11/18
ジャーナル フリーNitric oxide (NO) is synthesized from arginine by nitric oxide synthase, generating citrulline as another product, which can be recycled to arginine by argininosuccinate synthetase and argininosuccinate lyase. Rat argininosuccinate synthetase was expressed in Escherichia coli as a fusion protein with maltose binding protein, cleaved from the binding protein, and purified. The purified synthetase had no enzyme activity. Rat argininosuccinate lyase was expressed in E. coli using pET-3a as a vector, and purified. The purified enzyme had a specific enzyme activity of arginine formation of 2.6 μmol/min/mg protein at 37°C, the value being somewhat lower than those of the enzyme purified from various tissues. Antibodies against these enzymes were produced in rabbits. Immunoblot analyses showed that the two enzymes are most abundant in the liver, followed by kidney and testis. Smaller amounts of the enzyme proteins were present in other tissues. RNA blot analysis showed that the argininosuccinate synthetase mRNA was most abundant in the liver and kidney, followed by testis and other tissues. On the other hand, argininosuccinate lyase mRNA was most abundant in the testis, followed by kidney and liver, and by other tissues. These results show that argininosuccinate synthetase and argininosuccinate lyase are expressed both tissue-specifically and ubiquitously, and that practically all tissues have activities to convert citrulline to arginine.抄録全体を表示PDF形式でダウンロード (4015K) -
Masakazu Shiota, Fumiko Chinzei, Mitsuaki Moriyama, Kazuhiro Kimura, T ...1995 年 117 巻 5 号 p. 958-964
発行日: 1995年
公開日: 2008/11/18
ジャーナル フリーThe effects of increases in perfusion pressure and direction of perfusion flow (anterograde and retrograde) on hepatic carbohydrate metabolism were examined in the isolated perfused rat liver. Changing the direction of flow from anterograde (portal to caval vein) to retrograde (caval to portal vein) increased the rates of glycogenolysis and gluconeogenesis from sorbitol, lactate, pyruvate, and dihydroxyacetone. The extent of stimulation of gluconeogenesis by norepinephrine was higher during anterograde perfusion while stimulation by glucagon was higher during retrograde perfusion. Since the inflowing perfusion pressure was higher in retrograde perfusion (3.8 mmHg) than during anterograde perfusion (2.2 mmHg), we examined the effect of elevation in perfusion pressure on hepatic metabolism. In anterograde (portal to caval vein) perfusion, increases in perfusion pressure above the basal level (2.2 mmHg) caused increases in rates of glycogenolysis and gluconeogenesis with maximum rates at 4 mmHg. The extent of stimulation of gluconeogenesis by norepinephrine decreased and that by glucagon increased during perfusion at elevated pressure. At the same perfusion pressure (4 mmHg), there were no differences in rates of glycogenolysis and gluconeogenesis and in the responses to hormones between anterogradely and retrogradely perfused livers. The omission of Ca2+ ions from the perfusate abolished the effects of retrograde perfusion and of the elevation of perfusion pressure on carbohydrate metabolism. An infusion of A23187 (30 nM) induced an increase in both glycogenolysis and gluconeogenesis with unchanged perfusion pressure. The results suggest that elevated perfusion pressure during retrograde perfusion stimulates hepatic carbohydrate metabolism via a Ca2+-dependent process.抄録全体を表示PDF形式でダウンロード (813K) -
Satoshi Odo, Kei Kamino, Satoru Kanai, Tadashi Maruyama, Shigeaki Hara ...1995 年 117 巻 5 号 p. 965-973
発行日: 1995年
公開日: 2008/11/18
ジャーナル フリーA Ca2+-dependent lectin was purified from the hemolymph of a photosymbiotic bivalve, Tridacna derasa. An electrophoretically homogeneous form was obtained by using affinity chromatography with Sepharose 4B. More than 80% of the hemolymph protein was accounted for by this lectin. The apparent molecular mass of the lectin, in its native form exhibiting hemagglutinating activity, was estimated by gel filtration analysis to be approximately 480 kDa. On sodium dodecyl sulfate-polyacrylamide gel electrophoresis, in the absence of reductants, it migrated as a single band corresponding to a very large size, while in the presence of 2-mercaptoethanol, it migrated as two distinct bands of 23 and 46 kDa. These results indicate that the subunits were linked by disulfide bridges to form the native protein. After reducing pyridylethylation, each of the 23- and 46-kDa polypeptides was isolated by gel filtration in a mobile phase containing guanidine-HCI. The two polypeptides had the same amino-terminal sequence and a similar amino-acid composition, and in the presence of 2-mercaptoethanol gave a single band on isoelectric focusing at a pH of 6.0. The results suggested that the 46-kDa peptide is a homodimer of 23-kDa subunits held together by a covalent bond other than a disulfide linkage. This lectin required calcium ions for its activity. By ultraviolet spectrophotometry the association constant for the calcium ion was determined to be 0.88mM. The hemagglutinating activity decreased dramatically below pH 6.5, but re-increased to the original level when the solution was neutralized. Such a pH-dependent alteration of the ligand-binding activity was similar to that found in vertebrate asialoglycoprotein receptors. The amino-terminal sequence, determined to 29 residues, showed homology to some Call-dependent, C-type, animal lectins. This observation, together with the calcium ion requirement, suggests that the tridacnid lectin is a member of the C-type lectin family.抄録全体を表示PDF形式でダウンロード (1870K) -
Shin Murai, Toshiaki Arata, Akio Inoue1995 年 117 巻 5 号 p. 974-979
発行日: 1995年
公開日: 2008/11/18
ジャーナル フリーIt was shown by Miyanishi et al. [Miyanishi, T., Maita, T., Matsuda, G., and Tonomura, Y. (1982) J. Biochem. 91, 1845-1853] that the amino acid sequence around the reactive lysine residue is different between head B (P1-burst head) and head A of the myosin molecule. Thus, we synthesized these two peptides, and prepared rabbit polyclonal antibodies against them. Each antibody bound strongly with both peptides. However, the binding of the antibodies with S-1 was inhibited by the peptide used for the antigen but unaffected by the non-antigen peptide, suggesting that only antibodies specific to each head can bind with S-1. Myosin was absorbed by either antibody A or B, which was immobilized on protein A in Staphylococcus aureus cells. However, half of S-1 was absorbed by each of the antibodies. The S-1 prepared showed about 0.5 mol of initial P1-liberation per mol of S-1. The P1-burst size of S-1 unbound to the immobilized anti-A antibody increased to almost 1 mol/mol S-1, while that of S-1 unbound to the anti-B antibody decreased to 0.15 mol/mol S-1. These results suggest the existence of two kinds of heads in the myosin molecule.抄録全体を表示PDF形式でダウンロード (765K) -
Toshihide Okajima, Sachio Goto, Katsuyuki Tanizawa, Mitsuo Tagaya, Tos ...1995 年 117 巻 5 号 p. 980-986
発行日: 1995年
公開日: 2008/11/18
ジャーナル フリーA cDNA encoding porcine brain UMP-CMP kinase has been isolated using two oligonucleotide probes synthesized on the basis of the partial amino acid sequences of the purified enzyme. The isolated cDNA consisted of 1, 626 nucleotides including the coding region for a polypeptide of 196 amino acid residues with a calculated molecular weight of 22, 279. The enzyme showed an overall sequence identity of about 40 and 50%, respectively, with adenylate kinases from mammalian muscle and Escherichia coli and UMP-CMP kinases from Saccharomyces cerevisiae and Dictyostelium discoideum. The two highly conserved residues, Thr-39 and Leu-66, in adenylate kinases, which are located close to the adenine ring of the bound AMP, are replaced by Ala and Ile, respectively, at the corresponding positions in UMP-CMP kinases. The entire structural gene was inserted 3'-downstream of the strong promoter in the expression plasmid pET-3b. E. coli BL21(DE3) cells carrying the resultant plasmid produced the active enzyme in a soluble state, most efficiently upon induction at 37°C with 0.02mM isopropyl-β-D-thiogalactoside. The purified recombinant enzyme catalyzed specific phosphoryl transfer from ATP to UMP and CMP.抄録全体を表示PDF形式でダウンロード (2390K) -
Takahiko Kobayashi, Koichi Honke, Shinsei Gasa, Yoshihiro Saga, Tamots ...1995 年 117 巻 5 号 p. 987-992
発行日: 1995年
公開日: 2008/11/18
ジャーナル フリーBiosynthesis of sulfoglycolipid is markedly increased in a human renal cancer cell line, SMKT-R3. We investigated the sulfotransferase catalyzing the transfer of sulfate from 3'-phosphoadenosine 5'-phosphosulfate (PAPS) to gangliotriaosylceramide (Gg3Cer) in SMKT-R3 cells. On thin-layer chromatography, the reaction product comigrated with Gg3Cer III3-sulfate (SM2b), not with Gg3Cer II3-sulfate (SM2a). To examine which monosaccharide of Gg3Cer was sulfated, the product was treated with β-hexosaminidase A. Unlike authentic SM2a, of which the non-reducing terminal N-acetylgalactosamine was cleaved off, the product was not hydrolyzed. These results suggest that sulfate was transferred to the non-reducing terminal N-acetylgalactosamine. Gg3Cer sulfotransferase activity was independent of divalent cations but stimulated by Fe2+, and the optimal pH was approximately 6.5. The apparent Km values were 102 μM for PAPS and 348 umM for Gg3Cer. Gg3Cer II3, III3-bis-sulfate (SB2) and a sulfotransferase activity synthesizing SB2 from SM2a were also detected in SMKT-R3 cells.抄録全体を表示PDF形式でダウンロード (4064K) -
Hiroyuki Fukui, Hiroyuki Mizuguchi, Ye Qi Liu, Nai Ping Wang, Hideyuki ...1995 年 117 巻 5 号 p. 993-998
発行日: 1995年
公開日: 2008/11/18
ジャーナル フリーA protein having a high-affinity binding site for [3H]mepyramine (MBP) was purified to homogeneity from rat liver membranes. The purified MBP has a single type of binding site for [3H] mepyramine with Kd value of 18.5 nM, and its molecular weight was determined to be 56, 000 by SDS polyacrylamide gel electrophoresis. Amino acid sequences of twelve Cryptic peptides derived from MBP are highly homologous with those of rat debrisoquine 4-hydroxylase (cytochrome P450 2D1) and other rat P450 2D subfamily members. In immunoblotting analysis, an antibody against rat P450 2D1 stained a band corresponding to MBP with Mr of 56, 000; its migration position was clearly different from that of rat P450 2D1. Substrates and inhibitors of debrisoquine 4-hydroxylase potently displace [3H]-mepyramine binding to MBP. Quinine and quinidine showed 400 and 80 times, respectively, higher affinity for MBP than for debrisoquine 4-hydroxylase. These results suggest that MBP is a novel P450 2D family member.抄録全体を表示PDF形式でダウンロード (2876K) -
Toshihiro Fujii, Kei Yamana, Yoshiro Ogoma, Yoshiyuki Kondo1995 年 117 巻 5 号 p. 999-1003
発行日: 1995年
公開日: 2008/11/18
ジャーナル フリーThe interaction between chicken gizzard calponin and phospholipids was examined by sedimentation assay and affinity chromatography. Calponin was sedimented with phosphatidylserine (PS) and phosphatidylinositol (PI) vesicles but not with phosphatidylcholine (PC) vesicles. The apparent Kd values of calponin to PS and PI were calculated to be 1.3×106 and 1.5×106M-1, respectively. Domain mapping with chymotryptic digestion showed that the phospholipid-binding site resided within the N-terminal 22-kDa fragment, in which the bindings of actin, calmodulin, S100, and tropomyosin also occur. The amount of calponin bound to PS and PI vesicles decreased with increasing ionic strength or Ca2+ concentrations. The presence of MgCl2 was needed for the calponin-PS vesicle interaction. Calponin-binding proteins including actin, calmodulin, and S100 inhibited calponin binding to the phospholipid vesicles in a concentration-dependent manner, while tropomyosin had little effect on the binding. The inhibitory effects of calmodulin and S100 were found only in the presence of CaCl2. Neither caldesmon nor SM22 affected the binding.抄録全体を表示PDF形式でダウンロード (1962K) -
Tatsuya Izumi, Shingo Tajima, Takeji Nishikawa1995 年 117 巻 5 号 p. 1004-1007
発行日: 1995年
公開日: 2008/11/18
ジャーナル フリーLevels of type VI collagen mRNAs were determined in cultured skin fibroblasts derived from the upper, middle, and lower layers. The level of α1 (VI) collagen mRNA was the highest in the fibroblasts from the upper layer. In contrast, α2 (VI) collagen mRNA level in the fibroblasts from the lower layer predominated over those in the upper and middle layers. The level of α3 (VI) collagen mRNA was the same in the three dermal layers. When the fibroblasts from the upper layer were passaged successively, the α1 (VI) collagen mRNA level declined and α2 (VI) collagen mRNA level increased to those of the lower dermal fibroblasts. These results indicate that α1 (VI) and α2 (VI) collagen expressions in skin fibroblast cultures are heterogeneous and that successive passages of the upper dermal fibroblasts may result in transition of the phenotype of α1 (VI) and α2 (VI) expression to that of lower dermal fibroblasts. The modulation of the expression of α1 (VI) and α2 (VI) chains presumably reflects the process of cellular aging in vitro and should be a useful biochemical marker in studies of cell aging.抄録全体を表示PDF形式でダウンロード (2663K) -
Tsuneyoshi Funai, Arata Ichiyama1995 年 117 巻 5 号 p. 1008-1016
発行日: 1995年
公開日: 2008/11/18
ジャーナル フリーSerine:pyruvate/alanine:glyoxylate aminotransferase (SPT/AGT) of rat liver is localized in both mitochondria and peroxisomes. The rat SPT/AGT gene is single, but there are two species of mRNA which differ at their 5' termini due to transcription from two alternative initiation sites. The longer mRNA is translated from the first AUG codon and thereby directs synthesis of the 45 kDa precursor of mitochondrial SPT/AGT, which includes a mitochondria-targeting N-terminal signal sequence. Peroxisomal SPT/AGT is synthesized as a product of mature size (43 kDa) from the shorter mRNA, which starts 3' to the first AUG codon and thus is translated from a downstream AUG codon. In our previous immunocytochemical study, SPT/AGT was found to be localized only in peroxisomes, when a cDNA encoding 43 kDa SPT/AGT was expressed in COS cells. When a cDNA encoding the 45 kDa precursor was expressed, on the other hand, SPT/AGT was localized mostly in mitochondria, but a small number of peroxisomes were also positively stained [Yokota, S., Funai, T., and Ichiyama, A. (1991) Biomed. Res. 12, 53-59]. We show in this paper that 43 kDa SPT/AGT is also synthesized from the longer mRNA in an in vitro translation system through a leaky scanning mechanism. Although the first AUG initiator codon is in a suboptimal context, the amount of 43 kDa SPT/AGT synthesized from the longer mRNA was small, probably because a downstream stem-loop structure facilitates recognition of the first AUG initiator codon.抄録全体を表示PDF形式でダウンロード (4918K) -
Keiichi Fukuyama, Noritaka Ueki, Haruki Nakamura, Tomitake Tsukihara, ...1995 年 117 巻 5 号 p. 1017-1023
発行日: 1995年
公開日: 2008/11/18
ジャーナル フリーThe structure of plant-type [2Fe-2S] ferredoxin isolated from Spirulina platensis has been refined using diffraction data to 2.5 Å resolution by alternate cycles of simulated annealing and manual revision of the model. The final R factor is 19.9% for 2, 912 reflections with F>2σF, between 8.0 and 2.5 Å resolution. S. platensis ferredoxin, like other plant-type [2Fe-2S] ferredoxins, has a major a-helix flanking a sheet consisting of four β strands. The present refinement revises the conformation of residues 56-71, in which a one-turn helix was identified. Superposition of the Spirulina ferredoxin structure on the structures of other ferredoxins that have been well refined showed structural perturbation at a few residues on the amino and carboxyl termini and the turn between the first and second β-strands. The root-mean-square deviations of the corresponding Ca atoms of the pairs of ferredoxins range from 0.90 to 1.17 β for all the residues, but from 0.64 to 0.70 β if the few perturbed residues are excluded. Therefore, it may be concluded that the main-chain foldings of all the plant-type [2Fe-2S] ferredoxins are essentially the same. Electrostatic potential analysis showed that the molecular surface around the cluster is negatively charged, whereas that of the β-sheet of the other side is positively charged. The interaction between ferredoxin and ferredoxin-NADP+ reductase is discussed on the basis of the charge distributions of these molecules and biochemical data.抄録全体を表示PDF形式でダウンロード (1725K) -
Akiyoshi Tanaka, Harumi Fukada, Katsutada Takahashi1995 年 117 巻 5 号 p. 1024-1028
発行日: 1995年
公開日: 2008/11/18
ジャーナル フリーThermal unfolding of two forms of Aspergillus niger glucoamylase was observed by adiabatic differential scanning calorimetry (DSC) at pH 7. The DSC traces of the larger form of the enzyme, G1, and the shorter form which lacks the C-terminal starch-binding domain of G1, G2, could be resolved by assuming two-state unfolding of five and four independent components, respectively. The thermal unfolding of the starch-binding domain was found to be reversible, but that of the catalytic domain was irreversible. The DSC observation of G1 and G2 in the presence of β-cyclodextrin or 1-deoxynojirimycin showed that there was no domain interaction between the starch-binding domain and catalytic domain.抄録全体を表示PDF形式でダウンロード (652K) -
Satoru Harumiya, Akira Omori, Tsukasa Sugiura, Yuki Fukumoto, Hiroyuki ...1995 年 117 巻 5 号 p. 1029-1035
発行日: 1995年
公開日: 2008/11/18
ジャーナル フリーIn order to study the elastin-binding factors in blood, human plasma was applied to an α-elastin-Sepharose column. The column-binding fraction contained a 37-kDa protein, which was tentatively named EBP-37. Partial amino acid sequences of EBP-37 were determined. It had collagenous and non-collagenous domains. Homology searches of the sequences revealed that the protein is very similar but not identical to ficolins, transforming growth factor-β1 (TGF-β1)-binding proteins from porcine uterus membranes. Direct interaction of EBP-37 with elastin was confirmed by demonstrating the binding of the isolated EBP-37 to α-elastin on a nitrocellulose membrane using the EBP-37-specific antiserum. The existence of oligomers and multimers crosslinked by disulfide bonds was demonstrated by immunoblot analysis. Possible functions of EBP-37 are discussed.抄録全体を表示PDF形式でダウンロード (2674K) -
Michio Muguruma, So Nishimuta, Yoshito Tomisaka, Tatsumi Ito, Sueo Mat ...1995 年 117 巻 5 号 p. 1036-1042
発行日: 1995年
公開日: 2008/11/18
ジャーナル フリーTalin, a putative homodimer of 230-kDa polypeptides, was cleaved into the N-terminal 47-kDa and C-terminal 190-kDa fragments with calpain II. The 190-kDa fragment, but not the 47-kDa fragment, was found to bind to actin. The 190-kDa fragment possessed similar levels of activities to stimulate both polymerization of G-actin and α-actinin-dependent gelation of F-actin as did intact talin. Limited digestions of the 190-kDa fragment with chymotrypsin and papain resulted in partial and complete reductions, respectively, of both activities, although these digests contained 95- and 46-kDa major polypeptides, respectively, which were able to bind to actin. Whereas the 190-kDa fragment generated fully cross-linked oligomeric polypeptides on treatment with 1-ethyl-3 [3- (dimethylamino)-propyl] carbodiimide, the 95-kDa chymotryptic polypeptide generated heterologous polypeptides cross-linked partially with smaller polypeptides. The papain digest did not contain any cross-linkable polypeptide. Intact talin and the 47-kDa calpain fragment, but not the 190-kDa calpain fragment, were found to bind to phospholipid vesicles containing phosphatidylserine. These results indicate that the N-terminal and C-terminal domains play distinct roles in interacting with the membrane and cytoskeletal elements, respectively, and that the dimeric structure is also required for the latter interactions.抄録全体を表示PDF形式でダウンロード (3450K) -
Kwang-Pil Choi, Istvan Molnar, Mitsuo Yamashita, Yoshikatsu Murooka1995 年 117 巻 5 号 p. 1043-1049
発行日: 1995年
公開日: 2008/11/18
ジャーナル フリーThe 3-ketosteroid-Δ1-dehydrogenase (KS1DH) gene of Arthrobacter simplex IFO12069 cloned in Streptomyces lividans was overexpressed, resulting in production of the enzyme both extracellularly and intracellularly. The enzyme was purified by ammonium sulfate fractionation and chromatographies using DEAE-Toyopearl, Butyl-Toyopearl and Toyopearl HW55S from the supernatant of culture broth and cell-free extracts of S. lividans, and both preparations showed the same characteristics. The N-terminal amino acid sequence of both KS1DHs was M-D-W-A-E-E-Y-D, which coincided with the amino acid sequence deduced from the nucleotide sequence. Thus, the extracellular enzyme may derived from leakage of S. lividans cells during cultivation rather than secretion by processing of the signal sequence. The molecular weight of the enzyme was about 55, 000, identical with the size deduced from the nucleotide sequence (Mr 54, 329). The optimum conditions for its activity were pH 10.0 and 40°C. The enzyme catalyzed the conversion of several 3-ketosteroids, but those containing 11α-or 11β-hydroxyl group were converted at low rates. The amino acid sequence of KS1DH from A. simplex is similar to those of KS1DH of Pseudomonas testosteroni and fumarate reductase from Shewanella putrefaciens.抄録全体を表示PDF形式でダウンロード (1678K) -
Shoji Tajima, Hideki Tsuda, Nobunao Wakabayashi, Akira Asano, Shigeki ...1995 年 117 巻 5 号 p. 1050-1057
発行日: 1995年
公開日: 2008/11/18
ジャーナル フリーA 0.5 kb fragment of chicken DNA methyltransferase cDNA was PCR-amplified using a set of degenerate primers. A clone harboring a 5 kb insert was isolated from a cDNA library by screening with the PCR-amplified cDNA fragment as a probe. The elucidated nucleotide sequence gave a 4, 614 nucleotide open reading frame, and the predicted protein was highly homologous to the mouse and human DNA methyltransferases, especially in the amino acid sequence of the catalytic domain in the carboxyl-terminal region. The cysteine-rich region and Lys-Gly repeat first found in the mouse sequence were also conserved in chicken. However, about 250 amino acid residues in the amino-terminal portion of chicken DNA methyltransferase diverged from the amino-terminus of the mouse or human sequence. Northern blot analysis showed that the message of chicken DNA methyltransferase was expressed at high levels in the testis, in the lung and in Marek's virus-transformed chicken T-lymphoma cells. Expression of the chicken DNA methyltransferase in COS1 cells demonstrated that the enzyme is a so-called maintenance-type methylase. When poly(dGdC)-poly(dG-dC) was used as the methyl acceptor, to provide a measure of de novo methylase activity, the Km value for S-adenosyl L-methionine was about 5 μM, which was 10 times higher than that when poly(dI-dC)-poly(dI-dC) was used. The affinity of DNA methyltransferase for S-adenosyl L-methionine in catalyzing de novo-type methylation activity was lower than that in catalyzing maintenance-type activity, though it was still high enough for the enzyme to work as a de novo-type methylase under physiological conditions.抄録全体を表示PDF形式でダウンロード (4424K) -
Inshik Park, David H. Ives1995 年 117 巻 5 号 p. 1058-1061
発行日: 1995年
公開日: 2008/11/18
ジャーナル フリーInitial-rate kinetic measurements with the affinity purified 2-deoxyguanosine (dGuo) kinase from beef liver mitochondria yielded reciprocal plots which converged on the abscissa, with either dGuo or ATP as the varied substrate. The limiting Km, for dGuo was 4.7 μM, and that for ATP was 780 μM. One product, dGMP, was competitive with both substrates, while the other, ADP, was competitive with ATP and non-competitive with dGuo. Qualitatively identical results were obtained with an alternative substrate, dTTP, and with alternative product inhibitors, dIMP and dTDP. These results are consistent with a random Bi Bi kinetic mechanism, judging from the formation of a dead-end complex of the enzyme, dGuo and ADP. dGTP competes very strongly with ATP (K1=0.03 μM), but is non-competitive towards dGuo. The more weakly-bound dGDP is competitive with both substrates.抄録全体を表示PDF形式でダウンロード (454K) -
Shinobu Watarai, Yasunori Kushi, Rie Shigeto, Naoko Misawa, Yoshinobu ...1995 年 117 巻 5 号 p. 1062-1069
発行日: 1995年
公開日: 2008/11/18
ジャーナル フリーWe have established three kinds of monoclonal antibodies against gangliosides containing N-glycolylneuraminic acid (NeuGc) by immunization of BALB/c mice with the purified gangliosides inserted into liposomes comprising Salmonella minnesota R595 lipopolysaccharides, and fusion of spleen cells with a mouse myeloma cell line. One monoclonal antibody, SHS-1, which was generated by immunizing mice with purified i-active ganglioside(NeuGc), reacted specifically with the i-active ganglioside(NeuGc) used as an immunogen. Structurally related gangliosides, such as GM3(NeuGc), sialosylparagloboside (SPG) (NeuGc), or I-active ganglioside(NeuGc), corresponding gangliosides [GM3 containing N-acetylneuraminic acid (NeuAc), SPG(NeuAc), i-active ganglioside(NeuAc), and I-active ganglioside(NeuAc)], other gangliosides, or neutral glycosphingolipid (GSL) were not recognized by the monoclonal antibody. These findings indicate that the SHS-1 monoclonal antibody may be specific for NeuGc-containing i-active ganglioside. On the other hand, the other two monoclonal antibodies, MSG-1 and SPS-20, which were generated by immunizing mice with purified ganglioside GM3(NeuGc) and SPG(NeuGc), respectively, showed crossreactivity to structurally related gangliosides. The MSG-1 monoclonal antibody exhibited reactivity to ganglioside GM3(NeuAc). The SPS-20 monoclonal antibody also cross-reacted with SPG(NeuAc), i-active ganglioside(NeuGc), and i-active ganglioside(NeuAc). Neither MSG-1 nor SPS-20 reacted with corresponding gangliosides, other gangliosides, or neutral GSLs tested. Using the SHS-1 antibody specific for i-active ganglioside(NeuGc), we studied the expression of NeuGc-containing antigen in human colon cancer tissue. An NeuGc-containing glycoconjugate was detected in the colon cancer tissue.抄録全体を表示PDF形式でダウンロード (3660K) -
Takako Kitani, Sachiko Okuno, Hitoshi Fujisawa1995 年 117 巻 5 号 p. 1070-1075
発行日: 1995年
公開日: 2008/11/18
ジャーナル フリーThe activity of calmodulin-dependent protein kinase IV (CaM-kinase IV) was progressively decreased by incubation at 30°C with calmodulin in the presence of Ca2+, becoming one-half to one-fifth of the original activity within several minutes. The amount of calmodulin necessary to produce the maximal inactivation was approximately 1 mol for 1 mol of the enzyme. The inactivation of CaM-kinase IV by Ca2+/calmodulin was prevented by ATP in the presence of Mg2+, but such protection was not observed with either of the two alone or with a peptide substrate such as syntide-2. The activity of the calmodulininactivated enzyme was increased by incubation at 30°C with Mg2+ in the presence of EGTA, being completely restored to the original level within several minutes, indicating that the Ca2+/calmodulin-induced inactivation of the enzyme was not due to irreversible denaturation of the enzyme. Both the inactivation of CaM-kinase IV by Ca2+/calmodulin and the restoration of its activity by Mg2+/EGTA were time- and temperature-dependent reactions. Kinetic analysis revealed that the alterations of the enzyme activity were due mainly to changes in Vmax of the enzyme.抄録全体を表示PDF形式でダウンロード (739K) -
Yasuro Shinohara, Hiroyuki Sota, Fimi Kim, Mizuho Shimizu, Masanori Go ...1995 年 117 巻 5 号 p. 1076-1082
発行日: 1995年
公開日: 2008/11/18
ジャーナル フリーWe have developed a new method for analysis of the interaction between lectins and biotin-derivatized oligosaccharides involving a biosensor based on surface plasmon resonance. Complex type asialo-bi-, tri-, and tetra-antennary oligosaccharides were quantitatively converted into their biotin derivatives by incubating them with 6-(D-biotinyl)-aminohexanoyl hydrazide. This method was also applicable to sialyl sugar chains without any removal of sialic acid residues. The reaction mixture could be directly injected onto the streptavidin pre-immobilized surface of a sensor chip without any purification because of its fairly low reagent/carbohydrate molar ratio. The amounts of sugar chains required for interaction analysis by this method were as low as 1 pmol. The binding specificities of Sambucus sieboldiana lectin, Maackia amurensis lectin, Ricinus communis agglutinin-120 (RCA120), and concanavalin A could be rapidly determined qualitatively by this method. Furthermore, kinetic analysis of the interaction between RCA120, and complex type asialo-bi-, tri-, and tetra-antennary oligosaccharides revealed that both the association rate constant and the dissociation rate constant (kdiss) decreased with increasing numbers of terminal galactosyl residues. Because the tendency observed for kd, ss paralleled the elution order of these oligosaccharides on RCA120 immobilized affinity chromatography, kdiss might hold the key to determination of the elution order on affinity chromatography.抄録全体を表示PDF形式でダウンロード (827K) -
Hiroshi Kitagawa, Kazunori Tsuchida, Miho Ujikawa, Kazuyuki Sugahara1995 年 117 巻 5 号 p. 1083-1087
発行日: 1995年
公開日: 2008/11/18
ジャーナル フリーWe report the occurrence, in bovine serum of a soluble form, of N-acetylgalactosaminyltransferase activity, which is involved in chondroitin sulfate biosynthesis, and we describe a simple assay method for the enzyme. Chondroitin with the general structure [GlcAGalNAc]n was found to serve as acceptor substrates for the enzyme. To develop a routine assay, bovine serum was incubated with polymeric chondroitin and UDP- [3H] GalNAc. After the removal of labeled endogenous acceptors derived from serum by trichloroacetic acid treatment, the labeled chondroitin sulfate chains were separated from residual unincorporated label by centrifugation using syringe columns packed with Sephadex G-25, and the effluent fractions were monitored by scintillation counting. This simple method allowed us to perform multiple assays and to establish assay conditions for the serum N-acetylgalactosaminyltransferase involved in chondroitin polymerization.抄録全体を表示PDF形式でダウンロード (581K) -
Chikara Ebisui, Toshimasa Tsujinaka, Takashi Morimoto, Junya Fujita, A ...1995 年 117 巻 5 号 p. 1088-1094
発行日: 1995年
公開日: 2008/11/18
ジャーナル フリーC2C12 myoblasts fuse to form multinucleated myotubes and express muscle specific proteins during differentiation. To elucidate developmental regulation of intracellular proteolytic systems, enzymatic activities, protein and mRNA levels of proteasomes (20S and 26S) and lysosomal cathepsins (B, L, and H) were examined. Myoblasts were differentiated fully to myotubes 6 days after starting differentiation. In this developmental process, the 26S proteasome activity decreased, while the 20S proteasome activity increased. Expression of proteasome subunits of 20S (RC2, RC8) and regulatory components of 26S (S4, S7) was down-regulated, though total protein levels of proteasomes showed no remarkable changes. On the other hand, enzymatic activities of cathepsins B and B+L increased in association with an increase of their transcriptional and translational levels. Expression of their specific endogenous inhibitor, cystatin β, also increased. Maturation of the lysosomal proteolytic system was tightly linked to the differentiation process. These results suggested that signals for differentiation of myoblasts mediate a change of intracellular proteolytic systems, involving up-regulation of lysosomal cathepsins and downregulation of proteasomes.抄録全体を表示PDF形式でダウンロード (2315K) -
Mami Shoji-Kawaguchi, Shunji Izuta, Keiko Tamiya-Koizumi, Motoshi Suzu ...1995 年 117 巻 5 号 p. 1095-1099
発行日: 1995年
公開日: 2008/11/18
ジャーナル フリーWe studied the effects of various phospholipids on the DNA synthesizing reactions by calf thymus DNA polymerases α, δ, and ε. Of these three enzymes, DNA polymerase a was most sensitive to acidic phospholipids, i.e., phosphatidylinositol (PI), phosphatidylinositol 4-monophosphate (PIP), phosphatidylserine (PS), phosphatidic acid (PA), and cardiolipin (CAR). Of these acidic phospholipids, PI (from bovine liver) is of special interest because it inhibited DNA polymerase ε much more strongly than DNA polymerase α and δ. The inhibition of DNA polymerase ε by PI was competitive with the DNA template-primer and was noncompetitive with dTTP substrate. The K1 value was estimated to be 16 μM. These results indicate that PI from bovine liver can be used as a specific inhibitor for DNA polymerase s to analyze its role in DNA replication. Interestingly, the PI isolated from soybean, which has a different fatty acid composition, inhibited not only DNA polymerase ε but also DNA polymerase α.抄録全体を表示PDF形式でダウンロード (622K) -
Shwu-Bin Lin, Shyh-Shin Huang, Kong-Bung Choo, Pei-Jer Chen, Lo-Chun A ...1995 年 117 巻 5 号 p. 1100-1104
発行日: 1995年
公開日: 2008/11/18
ジャーナル フリーα-Fetoprotein (AFP) is a fetal protein which is absent in adult serum. However, the AFP gene is expressed in some neoplastic cells. According to the literature, AFP may play a role in accelerating the growth of cancer cells. In this report, 15meric antisense oligonucleotide analogues (phosphorothioates and methylphosphonates) and their chimeric forms, which were complementary to different regions of AFP mRNA, were synthesized, and their physical characteristics such as stability, melting temperature, and toxicity were compared. They were examined as to their inhibitory effects on the translation of AFP mRNA in a AFP-producing hepatoma cell line, HuH-7. We found that chimeric oligomers with methylphosphonate or phosphorothioate linkages at both the 5' and 3' ends were more effective than prototypic oligomers. Inhibition of 72% was achieved with a chimeric oligomer against the translational initiation region, at a concentration of 25 μM. No suppressive effect of the oligomers was observed on cell viability or albumin production, indicating the specificity of the inhibition.抄録全体を表示PDF形式でダウンロード (2841K) -
Takehisa Ishii, Maki Sato, Keiko Sudo, Masayuki Suzuki, Hiroshi Nakai, ...1995 年 117 巻 5 号 p. 1105-1112
発行日: 1995年
公開日: 2008/11/18
ジャーナル フリーThe effects of recombinant human hepatocyte growth factor (HGF) on liver growth and function of normal and partially hepatectomized rats have been examined. HGF was continuously administered into the jugular vein because it was rapidly eliminated from the plasma (t1/2α; _??_4.5min) and degraded. In normal rats, the labeling index of hepatocytes was increased about 6 times by the administration of HGF. HGF also decreased the prothrombin time and increased the hepaplastin and serum albumin content. In 70%-hepatectomized rats, HGF stimulated liver regeneration and increased the level of blood proteins such as hepaplastin in a dose-dependent manner. The stimulation of serum protein level seemed to result from not only the increase of hepatic cell number but also the direct effect of HGF on the protein production in hepatocytes, because HGF rapidly enhanced the protein synthesis prior to the increase of cell number and increased the mRNA content of albumin in the liver in vivo. In addition, a combination of heparin with HGF further accelerated the effects of HGF described above, possibly due to the decrease of HGF clearance. These findings suggest that HGF accelerates both the hepatic regeneration and function in vivo, and that rhHGF is clinically expected to be a potent therapeutic agent in hepatectomy and liver injury.抄録全体を表示PDF形式でダウンロード (2963K) -
Masayuki Ozawa1995 年 117 巻 5 号 p. 1113-1119
発行日: 1995年
公開日: 2008/11/18
ジャーナル フリーRecently, we described the isolation of a mouse cDNA clone encoding a novel Ca2+-binding protein, tentatively designated as reticulocalbin [Ozawa, M. and Muramatsu, T. (1993) J. Biol. Chem. 268, 699-705]. Reticulocalbin is a lumenal protein of the endoplasmic reticulum (ER) with a molecular weight of 44, 000 and has six repeats of a domain containing the high affinity EF-hand Ca2+-binding motif. The protein has the sequence, His-Asp-Glu-Leu (HDEL), at its carboxy terminus, which serves as a signal for its retention in the ER of cells. To examine the importance of the putative Ca2+-binding domains as well as the carboxyterminal HDEL sequence, we have cloned the human homologue of reticulocalbin. The sequence of this clone revealed a novel protein with 95% identity in amino acid sequence to the mouse reticulocalbin, indicating that this molecule has been evolutionarily conserved in mammals. As was found for the mouse reticulocalbin, the human homologue showed six repeats of a domain containing EF-hand motifs. Interestingly, conservation of the amino acid sequence was not restricted to the Ca2+-binding motifs, consistent with the possibility that reticulocalbin plays some role(s) besides Ca2+-binding. As was found for the mouse homologue, the protein has the HDEL sequence at its carboxy terminus instead of the Lys-Asp-Glu-Leu (KDEL) sequence, which is more common as a signal for the retention of resident proteins in the ER of animal cells. The conservation of the HDEL sequence in reticulocalbin in both species raises the possibility that this sequence has some roles in the function(s) of this protein family.抄録全体を表示PDF形式でダウンロード (2810K) -
Hiroyuki Inoue, Kenji Inagaki, Manabu Sugimoto, Nobuyoshi Esaki, Kenji ...1995 年 117 巻 5 号 p. 1120-1125
発行日: 1995年
公開日: 2008/11/18
ジャーナル フリーThe gene encoding L-methionine γ-lyase from Pseudomonas putida was cloned and the primary structure of the enzyme was deduced from its nucleotide sequence. The L-methionine γ-lyase gene was expressed in Escherichia coli. The amino acid sequences of BrCN-digested peptides agreed with the corresponding parts of the L-methionine γ-lyase sequence determined from the gene structure. The polypeptide is composed of 398 amino acid residues with a calculated molecular weight of 42, 626, corresponding to the subunit of the homotetrameric enzyme. The deduced amino acid sequence of L-methionine γ-lyase only showed extensive homology with other well known α, γ-elimination and/or γ-replacement pyridoxal 5'-phosphate-dependent enzymes, such as cystathionine γ-lyase, cystathionine γ-synthase, and O-acetylhomoserine O-acetylserine sulfhydrylase, that participate in the biosynthesis of sulfur amino acids. However, the deduced essential cysteine residue of L-methionine γ-lyase was not conserved in these enzymes. We confirmed the presence of a part of an open reading frame in the 3'-flanking region of the L-methionine γ-lyase gene, which showed high homology with the N-terminal region of pyruvate dehydrogenase (lipoamide) from E. coli, suggesting that it participates in the degradative pathway for L-methionine together with L-methionine γ-lyase.抄録全体を表示PDF形式でダウンロード (1917K) -
Tetsuya Uchikoba, Hiroo Yonezawa, Makoto Kaneda1995 年 117 巻 5 号 p. 1126-1130
発行日: 1995年
公開日: 2008/11/18
ジャーナル フリーCucumisin was isolated from prince melon sarcocarp by means of a simple purification procedure. Serine protease inhibitors such as soybean trypsin inhibitor, ovomucoid, and aprotinin had no effect on the enzyme activity. α2-Macroglobulin showed 38% inhibition of the original caseinolytic activity of cucumisin. The favorable synthetic substrates for cucumisin were Glt-Ala-Ala-Pro-Leu-pNA and Suc-Ala-Ala-Pro-Phe-pNA. The constant (kcat/Km) for Suc-Ala-Pro-Ala-pNA was found to be 30 times greater than that for Suc-Ala-Ala-Ala-pNA. The substrate specificity of cucumisin for oligopeptides and proteins was shown to be broad.抄録全体を表示PDF形式でダウンロード (1776K) -
Tetsu Saito, Shun-ichiro Kawabata, Takeshi Shigenaga, Yoko Takayenoki, ...1995 年 117 巻 5 号 p. 1131-1137
発行日: 1995年
公開日: 2008/11/18
ジャーナル フリーHemocytes of the horseshoe crab (limulus) contain a family of arthropodous peptide antibiotics, termed the tachyplesin family, and antibacterial protein, called anti-LPS factor, of which the former is located in the small (S) granules and the latter in the large (L) granules of the hemocytes. In our ongoing studies on granular components, we have identified here a novel defensin-like substance present in both L- and S-granules. This substance strongly inhibits the growth of Gram-negative and -positive bacteria, and fungi, such as Candida albicans. The isolated substance, tentatively termed “big defensin, ” consists of 79 amino acid residues, of which the COOH-terminal 37 residues have a sequence similar to those of mammalian neutrophil-derived defensins, especially rat defensin. Characterization of the disulfide motif in big defensin indicated that the disulfide array is identical to that of β-defensins from bovine neutrophils. One clear structural difference is that the limulus hemocyte-derived big defensin has an extension of the NH, -terminal hydrophobic sequence with 35 amino acid residues followed by the COOH-terminal cationic defensin portion. This amphipathic nature of big defensin seems likely to be associated with its potent antibacterial activity. Furthermore, antibacterial activities of the NH2-terminal hydrophobic region and the COOH-terminal defensin portion separated by tryptic digestion are significantly different: the former displays a more potent activity against Gram-positive bacteria, whereas the latter is more potent against Gram-negative bacteria. Big defensin, therefore, may prove to represent a new class of defensin family possessing two functional domains with different antimicrobial activities.抄録全体を表示PDF形式でダウンロード (3820K) -
1995 年 117 巻 5 号 p. 1138
発行日: 1995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 (58K)
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