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Volume 114 (1993)
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- Issue 4 Pages 449-
- Issue 3 Pages 299-
- Issue 2 Pages 163-
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Volume 114, Issue 2
Displaying 1-23 of 23 articles from this issue
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Valeria Vasta, Elisabetta Meacci, Marta Farnararo, Paola Bruni1993 Volume 114 Issue 2 Pages 163-166
Published: 1993
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSExogenous glutamine was metabolized by platelets to glutamate, aspartate, and CO2; whereas no lactate was formed. The amount of aspartate and CO2 produced from glutamine was reduced by the presence of glucose. On the other hand the rate of oxidation of glucose was reduced by the presence of glutamine. The potential energetic value of glutamine was lower than that of glucose, but the amino acid appeared to be a preferential respiratory substrate. The energy provision from glutamine also plays a role in the process of platelet activation, known to require an extra supply of ATP, since thrombin increased the rate of glutamine utilization in platelets. Upon cell activation an increase in aspartate and CO2 formation was observed and the stimulatory effect on glutamine oxidation by thrombin was also maintained in the presence of glucose.View full abstractDownload PDF (438K) -
Sachiko Okuno, Hitoshi Fujisawa1993 Volume 114 Issue 2 Pages 167-170
Published: 1993
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSCalmodulin-dependent protein kinase IV (CaM-kinase IV) is a Ca2+-responsive multifunc-tional protein kinase which occurs abundantly in the brain. When cDNA for rat brain CaM-kinase IV was expressed in Escherichia coli, the enzyme was produced in a good yield, but it did not show significant activity. The inactive recombinant CaM-kinase IV was phosphorylated and became highly active on incubation with a rat brain extract in the presence of both Ca2+/calmodulin and ATP/Mg2+. The recombinant CaM-kinase IV-activating activity in brain was one to two orders of magnitude higher than that in the other tissues examined. These observations suggest that CaM-kinase IV may undergo a posttranslational modification, probably Ca2+/calmodulin-dependent phosphorylation by CaM-kinase IV kinase, before exhibiting activity in the central nervous system.View full abstractDownload PDF (1926K) -
Masayoshi Jounouchi, Masatomo Maeda, Masamitsu Futai1993 Volume 114 Issue 2 Pages 171-176
Published: 1993
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSThe sequence (Gly-X-X-X-X-Gly-Lys-Thr/Ser) is conserved in nucleotide binding proteins including the α and β subunits of the ATP synthase. Various mutations were introduced in the αLys-175 and αThr-176 residues in the sequence (Gly-Asp-Arg-Gln-Thr-Gly-Lys-Thr, residues 169-176) of the Escherichia coli ATP synthase α subunit. Surprisingly, single amino acid substitutions drastically affected the subunit assembly of the enzyme. The entire enzyme assembly was lost by αLys-175→Phe (or Trp) or αThr-176→Phe (or Tyr) mutation. Other mutants had similar (αHis-175, αSer-175, αGly-175, αSer-176, and αHis-176 mutants) or lower (αAla-176, αCys-176, αLeu-176, and αVal-176 mutants) effects on assembly of the active enzyme compared with that of the wild-type. However, all these mutant enzymes except the αSer-176 enzyme showed enhanced cold sensitivities and reduced stabilities at high temperature. Mutant enzymes such as aGly-175 and αHis-176 showed low multi-site (steady state) catalysis, possibly due to loss of proper subunit-subunit interactions. These results suggest that the aLys-175 and αThr-176 residues are not absolutely essential for catalysis, but that they, or possibly the entire conserved sequence, are located in the key domain for the subunit-subunit interactions essential for enzyme stability and steady state activity. The roles of the α subunit residues are different from those of the corresponding β subunit residues (βLys-155 and βThr-156) which are essential for catalysis [Omote, H., Maeda, M., & Futai, M. (1992) J. Biol. Chem. 267, 20571-20576].View full abstractDownload PDF (1370K) -
Marika Eller, Jaak Järv, Reet Toomik, Ulf Ragnarsson, Pia Ekman, ...1993 Volume 114 Issue 2 Pages 177-180
Published: 1993
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSA set of stereoisomeric nonapeptides KRPSQRAKY with one, two, or all L-amino acid residues replaced by the corresponding D-amino acids, and two analogs with L- and D-threonine instead of serine, were synthesized and tested as substrates for protein kinase C. All of the peptides were phosphorylated by the enzyme. The maximal rate of the reaction with the all-D peptide was more than one order of magnitude lower than that for the all-L peptide with serine. The same applied to the peptides with D-Ser or with D-Arg in position +2 with respect to Ser. The Km values for the peptides containing one D-amino acid were close to that for the prototype peptide (53 μM). On the other hand, when two or more D-amino acids were present, the Km value increased considerably. Replacement of serine by threonine also reduced the phosphorylation rate and increased the Km values. One can conclude that the stereospecificity of protein kinase C is much less pronounced than that of protein kinase A, which is in agreement with the less clearly pronounced substrate specificity of the former enzyme.View full abstractDownload PDF (514K) -
Satoshi Watanabe, Koscak Maruyama1993 Volume 114 Issue 2 Pages 181-185
Published: 1993
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSDuring a survey of actin pointed end-capping protein in salt extracts of rabbit skeletal muscle, it was found that native tropomyosin (troponin-tropomyosin complex) inhibited the seed activity of β-actinin-capped actin filaments for actin polymerization. It turned out that β-actinin-capped actin fragments reannealed to form long filaments resulting in the decrease in seed numbers in the presence of troponin and tropomyosin. It appears that the reannealing of actin filaments was due to release of β-actinin from the actin filaments by troponin and tropomyosin. Either troponin or tropomyosin alone was not effective at all. Addition of an excess amount of β-actinin did not prevent the reannealing of β-actinin-capped actin filaments in the presence of troponin and tropomyosin.View full abstractDownload PDF (2563K) -
Ikuo Kawashima, Hideki Ozawa, Masaharu Kotani, Minoru Suzuki, Takehiro ...1993 Volume 114 Issue 2 Pages 186-193
Published: 1993
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSThe expression of N-glycolylneuraminic acid (NeuGc)-containing gangliosides in human melanoma cells grown both in culture and as xenografts in athymic (nu/nu) mice was analyzed extensively with specific mouse monoclonal antibodies (MAbs). Three MAbs (GMR8, GMR14, and GMR3) specific for GM3 (NeuGc), GM2 (NeuGc), and GD3 (NeuGc-NeuGc-), respectively, were used. Significant differences were observed in the ganglioside compositions between the cultured cells in vitro and the tumors grown in vivo. The major difference was that the cells cultured in serum-free medium did not express any NeuGc-containing gangliosides, whereas those grown in nude mice expressed a number of NeuGc-containing gangliosides, namely GM3 (NeuGc), GM2 (NeuGc), GD3 (NeuAc-NeuGc-), GD3 (NeuGc-NeuAc-), and GD3 (NeuGc-NeuGc-). The structures of these gangliosides were also determined chemically. No activity of CMP-NeuAc hydroxylase was demonstrated either in the melanoma cells cultured in vitro or in those grown in nude mice, suggesting that these cells incorporated NeuGc-containing glycoconjugates from the mouse sera and converted them to other NeuGc-containing gangliosides. The mouse sera contained only GM2 (NeuGc), but not the other NeuGc-containing gangliosides or any NeuAc-containing gangliosides.View full abstractDownload PDF (4376K) -
Hisako Kojima, Kazuya Hara, Rie Mineta-Kitajima, Fumiaki Taguchi, Shin ...1993 Volume 114 Issue 2 Pages 194-202
Published: 1993
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSWe have isolated and characterized a new subclonal cell line designated as MR 31, which was obtained by transfection of PC 12 cells with a glucocorticoid-regulated ras oncogene. The mRNA derived from the c-Ha-ras gene was proved to be expressed on exposure of the MR 31 cells to dexamethasone, the highest value being attained at 8h. MR 31 cells rapidly extended neurite-like processes within 24h in response to dexamethasone as well as nerve growth factor (NGF). The time of onset of neurite outgrowth induced by dexamethasone corresponded to the time when the highest ras mRNA level was observed. The cate-cholamine content of MR 31 cells was found to be twice that of PC 12 cells. A time course study on the effects of dexamethasone or NGF on cells showed that the former caused an increase in dopamine, a major catecholamine, to twofold the control level at 48h after the treatment, while the latter caused a decrease in the dopamine level. These effects on catecholamines were almost the same in MR 31 and PC 12 cells. The acetylcholinesterase activity of MR 31 cells was enhanced by both dexamethasone and NGF, whereas that of PC 12 cells was enhanced by NGF, but not by dexamethasone. The changes in acetylcholinesterase activity were correlated with neurite outgrowth. Electron-microscopically, MR 31 cells were not different from PC 12 cells. MR 31 cells exhibited extremely decreased tumorigenicity as compared with PC 12 cells. The morphological and biochemical properties of MR 31 cells remained constant, even after repeated passages. These findings indicate that MR 31 cells represent a stable transfectant exhibiting different properties from those of the parent PC 12 cells and are at the early stage of differentiating PC 12 cells, characterized by rapid response to NGF and dexamethasone.View full abstractDownload PDF (6112K) -
Taizo Suzuki, Masao Kawakita1993 Volume 114 Issue 2 Pages 203-209
Published: 1993
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSThe sarcoplasmic reticulum (SR) membranes of rabbit skeletal muscle were allowed to react with N-(3-pyrene)maleimide (PMI) at pH 7 at 30°C. The Ca2+-transporting activity of the SR membranes was reduced to 20% when PMI was bound to the extent of 1 mol/mol of Ca2+-transporting ATPase. The ATPase and the E-P forming activities were not affected by the binding of PMI up to 2mol/mol ATPase, indicating that PMI somehow uncoupled Ca2+-transport from ATP splitting. Permeability of the SR membranes to Ca2+ ions was increased in parallel with the loss of the Ca2+-transporting activity. Of several components of the SR membranes which are reactive with PMI, the ATPase protein was the only one whose modification by PMI was directly related to the loss of the Ca2+-transporting activity. Similar results were obtained with the light SR membrane fraction, which lacks the ryanodine receptor, a well-recognized Ca2+ channel. These results indicated that a Ca2+ channel that would have been latent or properly regulated in native ATPase somehow escaped from the normal control mechanism as a result of modification of its SH groups by PMI and went into runaway operation. The activated channel was specific for alkaline earth metal ions, so permeability to other solutes including Co2+, Ni2+, and sucrose remained unchanged after treatment with PMI. Permeability of the PMI-activated ion channel was regulated by Ca2+ in the external medium at concentrations which would saturate the Ca2+-transporting sites, suggesting a functional coupling between the selective gate for the Ca2+-transport and this channel, which may be the passageway of Ca2+ during the active transport.View full abstractDownload PDF (770K) -
Hirofumi Ohishi, Shinobu Fujii, Koji Tomoo, Toshimasa Ishida, Kiyoshi ...1993 Volume 114 Issue 2 Pages 210-214
Published: 1993
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSTo improve our understanding of why phospholipase A2 (PLA2) specifically catalyzes the hydrolysis of the fatty acid ester bond at position 2, not at position 1, of 1, 2-diacyl-3-sn-phosphoglycerides, the binding of each fatty acid chain of 1, 2-dilauroyl-L-phosphatidylethanolamine (DLPE), a natural substrate, to bovine pancreas PLA2 was examined by molecular dynamics (MD) simulations. Two different binding modes were considered, i.e., the respective hydrocarbon chains of 1- and 2-lauroyl fatty acid esters were located at the PLA2 binding sites usually observed in the complex crystals (Form A2) and at the reverse sites (Form A1). Although the total energies of both forms fluctuated within nearly the same range during the 80 ps MD simulations, the binding mode of DLPE to the PLA2 catalytic site through the coordination to Ca2+ was much more advantageous in Form A2 than that in Form A1; significant deviation of the Ca2+ position from its starting structure was observed in the MD simulation of Form A1. The result suggests the importance of Ca2+ in the selective recognition and catalytic function of PLA2 toward the 2-positioned fatty acid ester of phosphoglyceride substrates.View full abstractDownload PDF (445K) -
Kyosuke Sato, Yasuzo Nishina, Kiyoshi Shiga1993 Volume 114 Issue 2 Pages 215-222
Published: 1993
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSMammalian electron-transferring flavoprotein (ETF) has been reported to consist of two non-identical subunits and one FAD. The present paper shows that ETF purified from pig kidney contains one more molecule, an AMP. ETF was denatured by guanidine hydrochloride and ultrafiltered for the purpose of removing proteins. The filtrate was analyzed by reverse-phase chromatography. Two peaks appeared on the chromatogram: they were identified as FAD and AMP, and their molar amounts were identical, indicating that ETF contains one AMP molecule. ApoETF, which was prepared by KBr treatment of ETF, also contained one AMP molecule. These results clearly demonstrate that ETF has an AMP-binding site in addition to the FAD-binding site. AMP-free apoETF was prepared by guanidine treatment of ETF. Mixing AMP-free apoETF, FAD, and AMP produced recon-stituted ETF, which showed the same properties as native ETF. Mixing AMP-free apoETF and FAD produced AMP-free ETF, regardless of the coexistence of ATP or ADP: the AMP-binding site cannot bind FAD, ADP, or ATP. The enzymatic activity of the AMP-free ETF for electron transfer from substrate-reduced medium-chain acyl-CoA dehydrogenase to 2, 6-dichlorophenolindophenol was identical to that of native ETF. This indicates that the AMP contained in holoETF has no apparent influence on this enzymatic activity. A role of AMP recognized in this study is that AMP facilitates the formation of holoETF from AMP-free apoETF, FAD, and AMP.View full abstractDownload PDF (972K) -
Francisco J. G. Muriana, Valentina Ruiz-Gutierrez, M. Lourdes Gallardo ...1993 Volume 114 Issue 2 Pages 223-229
Published: 1993
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSThe lipid and fatty acid composition of the hepatopancreas and muscle of the prawn, Penaeus japonicus, were analyzed. The hepatopancreas was the main lipid storage organ, triglycerides and phospholipids being its major lipid components, while muscle contained mainly phospholipids. The fatty acid compositions of total lipids from the hepatopancreas and muscle were similar to those in other marine animals. The major fatty acids were palmitic (16:0), oleic (18:1 n-9), eicosapentaenoic (20:5 n-3), and docosahexaenoic (22:6 n-3) acids. The monoglycerides, diglycerides, triglycerides, and cholesterol esters from the hepatopancreas and muscle exhibited similar fatty acid patterns, but each lipid fraction was characterized by a specific paraffin chain composition. A blue carotenoprotein (λmax=640nm) containing astaxanthin was also extracted and purified from the hypodermis of the prawn. This blue carotenoprotein has a molecular weight of ca. 280, 000, which is much lower to those given for other crustaceans. The carotenoid prosthetic group was released from the carotenoprotein by the addition of acetone, and showed a hypsochromic shift to 470nm and the characteristic shape of free ketocarotenoids. TLC, infrared spectroscopy, chemical reduction, spectrophotometry, and qualitative analysis by HPLC were used to identify the astaxanthin as a unique chromophore group of the blue carotenoprotein. Moreover, HPLC studies suggested all- trans-astaxanthin to be the main component, which was accompanied by an epimer and its cis-isomer.View full abstractDownload PDF (763K) -
Eiichi Hoshino, Yasushi Sasaki, Mituo Okazaki, Kazutosi Nisizawa, Taka ...1993 Volume 114 Issue 2 Pages 230-235
Published: 1993
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSThe mode of action of two highly purified cellulases of exo- and endo-types from Irpex lacteus was investigated by using pure cellulosic substrates with different crystallinities derived from cellulose I. Exo-type cellulase saccharified all celluloses more effectively than endo-type enzyme, and the saccharification activities of both cellulases similarly increased with decreasing crystallinity of cellulose. The -DP-towering activity of exo-type cellulase remained similar for celluloses with higher crystallinity, while this cellulase showed a degradation mode resembling that of the endo-type enzyme for the substrates with lower crystallinity. Compared with exo-type cellulase, endo-type cellulase remarkably decreased the -DP of cellulose with higher crystallinity, while this activity was abated for cellulose with lower crystallinity. Thus, the effects of both cellulases became similar in the degradation of amorphous substrates such as H3PO4-treated cellulose. Endo-type cellulase produced several kinds of cellooligosaccharide from all kinds of cellulose used, while the product of the exo-type enzyme was only cellobiose from crystalline cellulose such as cotton and cotton linter even after a 12-h incubation period. The results indicate that each cellulase shows a typical mode of action (exo or endo) for crystalline cellulose, but that their characteristic modes of attack may change with decreasing crystallinity of cellulose.View full abstractDownload PDF (546K) -
Eiichi Hoshino, Yasushi Sasaki, Kazuo Mori, Mituo Okazaki, Kazutosi Ni ...1993 Volume 114 Issue 2 Pages 236-245
Published: 1993
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSThe interaction of two highly purified cellulases, exo- and endo-type cellulases from Irpex lacteus, with pure cotton and amorphous cellulose was investigated by electron microscopy. The morphological observations indicated that exo- and endo-type cellulases are both strongly adsorbed on the internal microfibril of cotton fiber before enzymatic hydrolysis, and then initiate their action toward the internal cellulose microfibrils with retention of the original shape. The two cellulases, however, caused considerably different morphological changes in cotton cellulose, and each cellulase seems to degrade native cellulose with a distinct mode of action. In the hydrolysis of cotton with exo-type cellulase, deep transverse cracks were produced and they extended from the fiber surface to the lumen structure located inside the fibers. In contrast, it was found that there were no deep cracks on fibers treated with endo-type cellulase, but severe internal erosion and cavitation occurred along fibril or microfibril layers inside the fibers. Thus, the degradation of cotton by exo- and endo-type cellulases yielded quite different morphological patterns, while little difference was found for regenerated celluloses. The mode of enzymatic hydrolysis of cellulose shown by cellulases with different degrees of randomness (exo and endo types) appears to be markedly affected by the fine structure of cellulose fibers.View full abstractDownload PDF (9710K) -
Mizue Morioka, Hiromichi Muraoka, Hajime Ishikawa1993 Volume 114 Issue 2 Pages 246-250
Published: 1993
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSSymbionin, a GroEL homologous molecular chaperone produced by an intracellular symbiont of the pea aphid, is autocatalytically phosphorylated in vitro at elevated temperatures. The phosphorylated symbionin showed a potent suppressive activity in spontaneous refolding of chemically denatured dihydrofolate reductase. When the 32P-labeled autophosphorylated symbionin was incubated with ADP, a portion of the radio-activity was transferred to ADP, suggesting that the autocatalytically phosphorylated symbionin contains high-energy phosphate bonds. It was also shown that when symbionin was incubated with [γ-32P] ATP and GDP, a large amount of the radioactivity was found in GTP, indicating that phosphate transfer between ATP and GDP is catalyzed by symbionin. These results suggested that in the endosymbiotic system symbionin functions as not only a molecular chaperone but also an energy-coupling protein.View full abstractDownload PDF (589K) -
Gazi Narun-Nahar Sultana, Yasuo Watanabe, Youichi Tamai1993 Volume 114 Issue 2 Pages 251-254
Published: 1993
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSThe translocation of orthophosphate, pyrophosphate, and polyphosphate into an organic phase was examined by means of the two-phase partition method. When the organic phase contained phosphatidylcholine (PtdCho), the phosphorus compounds translocated from the water-phase into the organic phase, depending on the concentrations of PtdCho and Ca2+ and the pH of the solution. Calcium ions were essential for the translocation. The concentration of Ca2+ necessary for the translocation appeared to depend on the molar concentration of the phosphorus compounds. Orthophosphate was translocated above pH 6.5, while pyrophosphate was translocated above pH 3.5. In contrast, polyphosphate was translocated in the whole pH range examined (pH 2.5-9.0), although approx. 9% of the total polyphos-phate remained in the water-phase even at alkaline pH. These results suggest that phosphorus compounds can interact with PtdCho when the phosphorus compounds have formed complexes with Ca2+.View full abstractDownload PDF (358K) -
Kazuya Yamanishi, Setsuko Yasuda, Yoshihiro Masui, Tsutomu Nishida, Yu ...1993 Volume 114 Issue 2 Pages 255-262
Published: 1993
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSThe structure of recombinant human carboxy-terminal-truncated macrophage colony-stimulating factor expressed in CHO cells was investigated. The bioactive protein ([-32-153] M-CSF), expressed from a nucleotide sequence that encoded a signal peptide of 32 amino acids and N-terminal amino acids numbers 1-153, was heterogeneous in terms of molecular mass, as analyzed by SDS-PAGE, because of the presence of N-linked sugar moieties. The primary structure of the polypeptide was determined by sequence analysis and amino acid analysis of the fragments obtained from lysylendopeptidase digests of reduced and alkylated M-CSF, and from pepsin digests of the intact molecule. A sugar chain was located only at Asn-122 of the two putative sites of N-glycosylation that were present per subunit. The homodimeric structure appeared to have seven disulfide bonds, formed by inter- or intra-molecular linkages, since there were no free thiol groups in the molecule. The assignment of disulfide bonds by sequence analysis using peptic fragments indicated the combinations of Cys7-Cys90, Cys48-Cys139, and Cysl02-Cysl46. Gel-filtration analysis of Ser31 [-32-153] M-CSF, in which the remaining Cys31 was replaced by Ser and which was expressed in COS cells, suggested that the mutein existed as a monomer. Our study shows that the disulfide-bond pairings of [-32-153] M-CSF that is expressed and post-translation-ally modified in mammalian cells are identical to those of Escherichia coli-derived [3-153]-M-CSF with only one intermolecular disulfide bond, namely, Cys3l-Cys3l.View full abstractDownload PDF (1986K) -
Yutaka Tsuneoka, Yoshinori Matsuo, Kazuhiko Iwahashi, Hiroaki Takeuchi ...1993 Volume 114 Issue 2 Pages 263-266
Published: 1993
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSGenetic polymorphism of the CYP2D6 gene [phenotypically individuals are either poor metabolizers (PM) or extensive metabolizers (EM)] has been reported to be associated with susceptibility to Parkinson's disease. We analyzed CYP2D6 genes from Japanese patients and controls, and found that EM/PM polymorphism is not a suitable marker for popula-tions with a low PM frequency. However, a novel mutant highly associated with Parkin-son's disease was discovered. The mutation was located at the HhaI site in exon 6 and changed a conserved amino acid residue, Arg296, to Cys296. The risk factor for the mutant homozygote was 5.56 (95% CI, 1.30-23.82). These results suggest that the HhaI polymor-phism in the CYP2D6 gene is a part of the molecular basis of Parkinson's disease.View full abstractDownload PDF (1463K) -
Susumu Y. Takahashi, Yoshimi Yamamoto, Yoshihiro Shionoya, Takashi Kag ...1993 Volume 114 Issue 2 Pages 267-272
Published: 1993
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSWhen an acid cysteine proteinase, which had been purified from the eggs of silkmoth, Bombyx mori, was incubated at pH 3.6, enzymatic activity appeared after a few minutes, lag period, indicating that the purified cysteine proteinase was a latent form. SDS-poly-acrylamide gel electrophoresis showed that after the incubation the latent form of the enzyme (47 kDa) disappeared and the active (39 kDa) form of the enzyme appeared, suggesting that the latent form was processed to the active form under acidic conditions (pH 3.6). The NH2-terminal 22-residue sequence of the active form was determined. The conversion of the latent form to the active form was completely blocked by E-64, which is a specific inhibitor of cysteine proteinases. The results strongly suggest that the processing might be autocatalytic. The latent form (47 kDa) disappeared in the silkmoth eggs during embryonic development and concomitantly with its disappearance, the 39-kDa form appeared, indicating that in vivo the enzyme is activated in a similar manner to that observed in in vitro experiments.View full abstractDownload PDF (5321K) -
Shiro Matsuura, Yonekazu Hamano, Hiroshi Kita, Yutaka Takagaki1993 Volume 114 Issue 2 Pages 273-278
Published: 1993
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSSeven of 20 mouse monoclonal antibodies to OA, OA 8-2, OA 10-8, OA 22-22, OA 227-11, OA 296-1, OA 423-3, and OA 958-2, were studied as to their binding to OA in organic solvents. OA 423-3 (IgG1-x) and OA 958-2 (IgG1-x) in 90-100% methanol retained their binding activities with both immobilized and free antibodies. Whereas OA 8-2 (IgG2a-x), OA 10-8 (IgG1-x), OA 22-22 (IgG2a-x), OA 227-11(IgG1-x), and OA 296-1(IgM-x) did not bind to OA in over 50-60% methanol. The results of a non-competitive inhibition assay for OA indicated that in a methanolic or ethanolic solution, the binding ability of immobilized OA 423-3 decreased as the concentration of each alcohol increased. The concentration of OA at the midpoint between the upper and lower plateaus of the inhibition curve was 0.18 ng/ml in 0% methanol and 570 ng/ml in 100%, respectively. In 0-50% of each of acetone, diethyl ether, and benzene in methanol, the binding ability of OA 423-3 remained at the level in 100% methanol. OA 958-2 showed similar binding properties to OA 423-3. No relationship between the subclass of the immunoglobulin and the binding activity of the antibody in organic solvents was observed. These results indicate that the OA 423-3 and OA 958-2 antibodies are useful for the development of a new ELISA method for OA in organic solvents.View full abstractDownload PDF (623K) -
Yuan Lin, Ryoki Ishikawa, Kazuhiro Kohama1993 Volume 114 Issue 2 Pages 279-283
Published: 1993
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSWe have previously shown that caldesmon at low concentrations stimulates the interaction between actin, myosin, and ATP, while at high concentrations it inhibits the interaction [Ishikawa, R., Okagaki, T., Higashi-Fujime, S., & Kohama, K. (1991) J. Biol. Chem. 266, 21784-21790]. When the effect of caldesmon at low concentrations was monitored by measuring myosin ATPase activity in the absence of actin, the effect was slightly but significantly stimulatory; and at higher concentrations no inhibitory effect was observed. Therefore, we related the stimulatory effect with the myosin-binding property of caldesmon. In the presence of actin, a low concentration of caldesmon was not enough to evince the stimulatory effect: myosin concentration must also be low. This is because the stimulatory effect was obscured when myosin concentration was elevated. Ca2+-calmodulin abolished the stimulatory effect of caldesmon. However, the concentration of calmodulin required to abolish the stimulation was higher than that required to abolish the inhibition.View full abstractDownload PDF (493K) -
José-Manuel Martin-Alonso, Sikha Ghosh, Miguel Coca-Prados1993 Volume 114 Issue 2 Pages 284-291
Published: 1993
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSIn the anterior segment of the mammalian eye, the ocular ciliary epithelium produces the aqueous humor, a fluid that nourishes and protects the avascular tissues from oxidative stress. This report details the results of a study of molecular cloning, sequencing, and expression of plasma glutathione peroxidase (GPx-P) from the bovine ocular ciliary epithelium. The bovine GPx-P cDNA contains an open reading frame of 226 amino acids with a calculated molecular weight of 24, 860. The corresponding amino acid sequence showed an overall identity of 88% with the human GPx-P, 88.5% with the rat GPx-P, and 46.4% with the cellular bovine glutathione peroxidase (GPx-1). The levels of GPx-P and GPx-1 transcripts in ocular tissues were analyzed and the ciliary epithelium was found to express the highest levels of GPx-P transcripts in human and bovine eyes, whereas the cornea of calf eyes expressed the highest levels of GPx-1 transcripts. Surprisingly, the lens, on which oxidants have profound effects leading to cataract formation, expressed the lowest levels of GPx-P and GPx-1 transcripts in human donor eyes. These results provide new evidence of differential gene expression of the GPx-P and GPx-1 forms in the mammalian eye and stresses the functional role of the ocular ciliary epithelium in protecting the anterior segment of the eye from oxidative damage.View full abstractDownload PDF (2648K) -
Satoshi Suzuki, Kohnosuke Mitani, Katsuhiro Kuwabara, Yoshiaki Takahas ...1993 Volume 114 Issue 2 Pages 292-296
Published: 1993
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSPc-1 and Pc-2 hypervariable minisatellites of the mouse show a structural similarity and hence have been compared in terms of chromosomal location and mutability. Polymor-phisms of the Pc-2 locus detected by analyses of Southern blotting and polymerase chain reaction were used for typing 61 backcross mice that were obtained by mating (C57BL/6×MSM)F1 females to MSM males. Segregation patterns indicated that the Pc-2 locus maps to the 60 cM region of chromosome 6, while that of Pc-1 maps to chromosome 4. Simultaneous mutation of the two loci was examined in 101 F1 mice. Pc-1 and Pc-2 loci showed mutation rates of 14.4 and 5.0% per gamete, respectively. Of the nine mice with a Pc-2 mutation, four carried a Pc-1 mutation at the same maternally- or paternally-derived allele. The rate of comutation was 44% (4/9), higher than the 12.2% (24/197) of Pc-1 mutation alone. This suggests that the germline mutation of the two hypervariable minisatellites may result from a common mechanism.View full abstractDownload PDF (2859K) -
1993 Volume 114 Issue 2 Pages 297
Published: 1993
Released on J-STAGE: November 18, 2008
JOURNAL FREE ACCESSDownload PDF (42K)
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