巻号一覧
100 巻, 2 号
選択された号の論文の32件中1～32を表示しています
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Hiromi TAKANO-OHMURO, Kazuhiro KOHAMA1986 年 100 巻 2 号 p. 259-268
発行日: 1986年
公開日: 2008/11/18
ジャーナル フリーPhosphorylation of the 20, 000 Mr light chain (L20) of gizzard myosin reversibly increased the mobility of myosin in pyrophosphate polyacrylamide gel electrophoresis (PP1 PAGE). Gizzard heavy meromyosin (HMM) with phosphorylated L20 also moved faster than that with unphosphorylated L20. This mobility increase of HMM is large enough to account for that of intact myosin.
Scallop myosin, desensitized by removing its regulatory light chain, was combined with L20 and subjected to PP1 PAGE. Hybrid myosin with the phosphorylated light chain moved faster than that with the unphosphorylated light chain. No such effect of light chain phosphorylation was observed with phosphorylatable light chain from breast or ventricular myosin.
Thus, gizzard, but not breast or ventricular phosphorylatable light chain is furnished with the ‘regulatory’ property that its phosphorylation increases myosin mobility in PP1 PAGE.抄録全体を表示PDF形式でダウンロード (4939K) -
Kunio MIKI, Yasuaki II, Motomu YUKAWA, Akira OWATARI, Yukinori HATO, S ...1986 年 100 巻 2 号 p. 269-276
発行日: 1986年
公開日: 2008/11/18
ジャーナル フリーThe crystal structures of sperm whale metmyoglobins reconstituted with four modified hemes, isopemptoheme, pemptoheme, 2-ethyldeuteroheme, and 4-ethyldeuteroheme, have been determined and refined at 2.2Å resolution to R=0.217, 0.218, 0.213, and 0.222, respectively. All the crystals of these myoglobins are isomorphous with that of native metmyoglobin. The structural changes of the modified myoglobin from the native myoglobin were examined on difference Fourier maps; the orientation of 4-ethyldeuteroheme in the heme pocket is such that the heme is rotated by 180° about an axis through the α-γ-meso carbons, whereas the orientations of the other three hemes are the same as that of the protoheme in the native myoglobin. The changes of the structures around the heme become greater in the order of isopemptoheme, 2-ethyldeuteroheme <pemptoheme <4-ethyldeuteroheme. The magnitudes of the changes seem to be related to the oxygen affinities of these four reconstituted myoglobins.抄録全体を表示PDF形式でダウンロード (963K) -
Kunio MIKI, Shigeharu HARADA, Yukinori HATO, Seigo IBA, Yasushi KAI, N ...1986 年 100 巻 2 号 p. 277-284
発行日: 1986年
公開日: 2008/11/18
ジャーナル フリーThe crystal structures of sperm whale metmyoglobins reconstituted with three kinds of modified heroes, 2, 4-diisopropyldeuteroheme, 2-isopropyl-4-vinyldeuteroheme, and 2-vinyl-4-isopropyldeuteroheme, have been determined and refined at 2.2Å resolution to R=0.216, 0.219, and 0.195, respectively. All the crystals of these myoglobins are isomorphous with that of native metmyoglobin. The 2-vinyl-4-isopropyldeuteroheme was found to be in a reverse orientation, in which the heme plane is rotated by 180° about an axis through the α-γ-meso carbons, whereas the orientations of the other two hemes were the same as that of protoheme in native myoglobin. In the myoglobins with 2, 4-diisopropyldeuteroheme and 2-vinyl-4-isopropyldeuteroheme, both of which have lower oxygen affinities than native myoglobin, the bulky isopropyl side chain pushes Phe 43 0.7Å toward His 64 (the distal histidine) in the former, and the whole E helix at most 1.5Å, including a 0.7Å shift of the His 64 imidazole ring, in the latter. The changes of the structures prevent His 64 from forming a hydrogen bond with the liganded oxygen molecule, so that these two modified myoglobins show low oxygen affinities. On the other hand, there is no such drastic displacement in myoglobin with 2-isopropyl-4-vinyldeutero-heme, which has a slightly higher oxygen affinity than native myoglobin.抄録全体を表示PDF形式でダウンロード (881K) -
Daisaku SATOMI1986 年 100 巻 2 号 p. 285-292
発行日: 1986年
公開日: 2008/11/18
ジャーナル フリーThe quantitative changes and metabolism of GABA and putative amino acid neurotransmitters during early developmental stages in the organotypic culture of newborn mouse cerebellum were examined by using the high-performance liquid chromatograph (HPLC) technique. D-[U-14C]Glucose was used as a precursor of amino acids.
To analyze amino acid neurotransmitters, explants were incubated for 4 weeks under standard conditions. The amount of GABA linearly increased from 8.7±1.3nmol/mg protein (2days in vitro, 2 DIV) to 26.5±6.1nmol/mg protein (15 DIV) and was saturated after that (24.0±3.6nmol/mg protein at 30 DIV). During the period of GABA increase, the capability for GABA synthesis from [14C]glucose increased rapidly from 3.03±0.67nCi/mg protein (2 DIV, 3h incubation) to 9.32±1.34nCi/mg protein (15 DIV, 3h incubation).
In the case of glutamic acid, a putative neurotransmitter of granule cell parallel fibers in the cerebellum, the amount in explants was nearly constant during incubation, in contrast with the fact that the amount in vivo gradually increased. However, the capability for glutamic acid synthesis from [14C]glucose increased from 10.80±3.01nCi/mg protein (2 DIV, 1h incubation) to 27.62±4.71nCi/mg protein (22 DIV, 1h incubation). In the case of taurine, found in abundance in fetal brain and supposed to play a specific role in the development and maturation of the central nervous system, the amount in explants decreased from 139.8±4.0nmol/mg protein (2 DIV) to 54.0±0.8nmol/mg protein (30 DIV). The pattern of change of taurine was similar to that observed in vivo.
On considering together the results obtained in this report and the previous one (Satomi, D. (1983) J. Biochem. 94, 785-791), it was presumed that in the organotypic culture of newborn mouse cerebellum, biochemical changes of neurotransmitters are closely related to morphological development in nearly the same manner as in vivo.抄録全体を表示PDF形式でダウンロード (1249K) -
Khalid Majid FAZILI, Mohammad Abul QASIM1986 年 100 巻 2 号 p. 293-299
発行日: 1986年
公開日: 2008/11/18
ジャーナル フリーBuffalo liver cathepsin B was isolated by acid extraction, ammonium sulfate fractionation, Sephadex gel filtration, DEAE-Sephadex chromatography and Sephacryl S-300 chromatography. The enzyme preparation was found to be homogeneous by gel filtration and SDS-polyacrylamide gel electrophoresis but could be resolved into two major and four minor protein bands on polyacrylamide gel electrophoresis in the absence of SDS. The enzyme showed catheptic activity against synthetic substrates such as BANA and BAPNA as well as against denatured hemoglobin. Various physico-chemical and enzymatic properties of the enzyme, such as molecular weight, Stokes radius, frictional coefficient, pH optimum, Michaelis constant, and Vmax, were determined. The values of these parameters were 27, 500, 2.41nm, 1.2, 6.5, 2.08mM, and 42.4units/mg, respectively. The hydrodynamic properties suggest a compact and globular conformation for this enzyme. Various compounds were tested for their influence on the activity of cathepsin B. Of these compounds, membrane phospholipids were found to increase significantly the activity of this enzyme. This increase in activity could be of physiological importance since the concentration of phospholipids is increased after endocytosis and autophagy.抄録全体を表示PDF形式でダウンロード (1161K) -
Takashi HASHIMOTO, Tonao KUWABARA, Nobuteru USUDA, Tetsuji NAGATA1986 年 100 巻 2 号 p. 301-310
発行日: 1986年
公開日: 2008/11/18
ジャーナル フリーPeroxisomes were obtained by sucrose density gradient centrifugation from the livers of di(2-ethylhexyl)phthalate-fed rats, and the membranes were prepared by carbonate extraction (Fujiki, Y., Fowler, S., Shio, H., Hubbard, A.L., & Lazarow, P. B. (1982) J. Cell Biol. 93, 103-110). The integrated membrane polypeptides were solubilized with sodium dodecyl sulfate, and purified by repeated polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate. Separation of 70 and 68kDa polypeptides was not attempted in the present study because of their close migration in polyacrylamide gel electrophoresis. Other polypeptides with apparent molecular masses of 41, 27, 26, and 22kDa were purified to near homogeneity. Antibodies were raised against these purified preparations. The 68kDa polypeptide is suggested to be produced by the proteolytic modification of 70kDa polypeptide, since the former increased concomitantly with decrease of the latter when the liver homogenate was incubated, and this change was prevented in the presence of leupeptin during the incubation. The 41kDa polypeptide was a minor component. The 70 and 68kDa polypeptides and 41kDa polypeptide and their antibodies were cross-reactive, but the relation of these polypeptides was not clear. The 27 and 26kDa polypeptides seemed to be another species of membrane polypeptides, although the relationship of these two polypeptides remains to be clarified. The 22kDa polypeptide is not related to other membrane polypeptides.
The results of immunoblot analysis of subcellular fractions of the liver and an electron microscopic immunocytochemical study to locate the antigenic sites with protein A-gold complex suggest that all of these polypeptides are localized on peroxisomal membranes.
On proliferation of rat liver peroxisomes by administration of di(2-ethylhexyl)-phthalate, a peroxisome proliferator, all of these polypeptides were markedly in-creased.抄録全体を表示PDF形式でダウンロード (4399K) -
Mari OGATA, Tatsuhiko YAGI1986 年 100 巻 2 号 p. 311-318
発行日: 1986年
公開日: 2008/11/18
ジャーナル フリーPyruvate dehydrogenase from DesulJovibrio vulgaris Miyazaki F was partially purified from the soluble fraction of the bacterial sonicate, and characterized. The enzyme catalyzes oxidative decarboxylation of pyruvate to produce acetyl-CoA, in contrast to statements in current review articles in which acetyl phosphate is indicated to be a direct decomposition product of pyruvate in sulfate-reducing bacteria. The established reaction stoichiometry is: pyruvate+CoA+FMN→acetyl-CoA+CO2+FMNH2. The Km values are 2.9mM for pyruvate, 32μm for CoA and 6.7μmol for FMN. Participation of thiamine diphosphate in the enzymic process was not proven. 2-Oxobutyrate, but not 2-oxoglutarate, can substitute for pyruvate. The three flavin compounds, FMN, FAD, and flavodoxin, as well as clostridial ferredoxin, serve as electron carriers for the enzyme. Thus the enzyme is a kind of pyruvate synthase [EC 1.2.7.1], but acts in the direction of pyruvate degradation in the growing cells. The rate of cytochrome c3 reduction is extremely low, but in the presence of flavodoxin as an electron mediator, the reduction rate of cytochrome c3 becomes faster than the reduction rate of flavodoxin alone. It seems that the physiological electron acceptor for this enzyme is flavodoxin, which might be complexed with cytochrome c3 to produce a very efficient electron transfer system in the cell. The soluble fraction of D. vulgaris cells has been proved to contain, in addition to the pyruvate dehydrogenase, lactate dehydrogenase (Ogata, M., Arihara, K., & Yagi, T. (1981) J. Bioehem. 89, 1423-1431), phosphate acetyltransferase and acetate kinase, i.e., all the enzymes necessary to convert lactate to acetate, producing ATP by substrate level phosphorylation.抄録全体を表示PDF形式でダウンロード (641K) -
Rikurou HAYASHI, Shoji TAJIMA, Akira YAMAMOTO1986 年 100 巻 2 号 p. 319-331
発行日: 1986年
公開日: 2008/11/18
ジャーナル フリーLipoprotein lipase [EC 3.1.1.34, LpL] was purified from human postheparin plasma (PHP) almost to homogeneity (a 210, 000-fold purification) using columns of heparin-Sepharose, hydroxylapatite, and concanavalin A-Sepharose, and its properties were compared with the purified bovine milk LpL. The specific activity of the PHP-LpL was 26mmol free fatty acids (FFA)/h/mg of protein at 37°C; close to that of bovine milk LpL (35mmol FFA/h/mg). For both enzyme preparations, the pH optimum (about 8.7) and the inhibition by sodium chloride were almost the same. The apparent Michaelis constants were also similar; 2.5mM for human PHP-LpL and 2.1mM for bovine milk LpL. The apparent molecular weight of the purified human PHP-LpL was 58, 000 by polyacrylamide gel electrophoresis in the presence of sodium dodecylsulfate, slightly larger than that of the bovine milk LpL (56, 000). Although the amino acid composition of the two LpL preparations had only slight differences, antibody raised against bovine milk LpL cross-reacted very weakly with purified human PHP-LpL. With 1% bovine serum albumin, bovine milk LpL was highly stable, but the human PHP-LpL was unstable; it lost 60% of its activity within 60min at 0°C. In the absence of apolipoprotein C-II (apo C-II), the activity of human PHP-LpL was very weak. However, human PHP-LpL was activated by apo C-II more strongly than bovine milk LpL; the fold activation of human PHP-LpL by apo C-II was 7-8 times that of bovine milk LpL. The apparent Km value of human PHP-LpL for apo C-II (1.00±0.58μM) was larger than that of bovine milk LpL (0.15±0.03μM).抄録全体を表示PDF形式でダウンロード (1949K) -
Yusei YAMAMOTO, Takafumi OHMURA, Kazuyoshi KAWAKAMI, Kaoru ONOUE, Hiro ...1986 年 100 巻 2 号 p. 333-340
発行日: 1986年
公開日: 2008/11/18
ジャーナル フリーPhytohemagglutinin (PHA) and a tumor-promoting phorbol ester, 12-O-tetrade-canoylphorbol-13-acetate (TPA) act synergistically to induce interleukin 2 (IL2) mRNA in human lymphocytes in vitro. The induction was inhibited by a potent inhibitor of protein kinase C (C-kinase), 1-(5-isoquinolinylsulfonyl)-2-methylpiperazine (H-7) at less than 10μM. H-7 inhibited C-kinase activity itself in lymphocytes at the same range of the concentration but did not interfere with the translocation of C-kinase from the cytosol to the membrane fraction of the lymphocytes induced by TPA. H-7 is also known to inhibit cAMP-dependent protein kinase (A-kinase) and cGMP-dependent protein kinase (G-kinase). However, the lymphocytes cultured with dibutyryl cAMP or dibutyryl cGMP could not be activated to produce IL2 mRNA. These results show that activation of C-kinase but not A-kinase and G-kinase is necessary in signal transduction for IL2 gene expression. Prostaglandin E2, which is known to elevate intracellular cAMP level, also inhibited IL2 mRNA induction in the lymphocytes stimulated with PHA and TPA. Addition of α-methylornithine and methylglyoxal his (guanyl hydrazone), which inhibit polyamine synthesis, did not affect the induction of IL2 mRNA in the lymphocytes stimulated with PHA and TPA, indicating that polyamine synthesis is not necessary for IL2 mRNA induction.抄録全体を表示PDF形式でダウンロード (1501K) -
Hitoshi UEDA, Yoshiaki HAGINO, Mayumi ONO, Michihiko KUWANO1986 年 100 巻 2 号 p. 341-348
発行日: 1986年
公開日: 2008/11/18
ジャーナル フリーWe have recently isolated retinoic acid-resistant clones U-2 and U-3 from human breast cancer cell line MCF-7 (Ueda et al. (1985) Cancer Res. 45, 3332-3338). Growth of MCF-7 cells was found to be stimulated by estradiol but that of U-2 or U-3 was not. Cytosol from U-2 or U-3 cells contained no detectable estradiol receptor activity, whereas that from the parental MCF-7 cells showed estradiol receptor activity of 32fmol/mg cytosol protein with a Kd of 2.6×10-10M by Scat-chard analysis. Sucrose gradient centrifugation analysis of the cytosol fraction confirmed the presence of estradiol receptor activity in MCF-7 but not in U-2. Cytosol from MCF-7 and U-2 cells showed progesterone receptor activities of 106fmol/mg protein with a Kd of 7.4×10-10M and 13fmol/mg protein with a Kd of 9.9×10-10M, respectively. Addition of estradiol to the culture medium of the cells increased the level of progesterone receptor about 2-fold in MCF-7, but not in U-2. U-2 or U-3 cells showed about 5-fold higher resistance to an antiestrogen, tamoxifen, than MCF-7, and they were also 300- to 1, 000-fold more resistant to other anti-estrogens, epitiostanol and medroxyprogesterone, than MCF-7. The altered cellular sensitivity of U-2 or U-3 to the hormone antagonists is discussed in relation to the absence or presence of hormone receptors.抄録全体を表示PDF形式でダウンロード (563K) -
Shobha CHAR, Karumathil P. GOPINATHAN1986 年 100 巻 2 号 p. 349-357
発行日: 1986年
公開日: 2008/11/18
ジャーナル フリーArginyl-tRNA synthetase [L-Arg: tRNAArg ligase (AMP forming) EC 6.1.1.19] has been purified to homogeneity from Mycobacterium sinegmatis SN2. The enzyme is a monomer of molecular weight 56, 000. The kinetic patterns obtained by initial velocity and product inhibition studies are consistent with a rapid equilibrium random ter ter mechanism. Polyamines stimulated the formation of arginvl-tRNA, the stimulation being more significant at sub-optimal Mg2+ concentrations. Initial velocity studies performed in the presence of sub-optimal Mg2+ and spermine also indicated that the kinetic mechanism remained sequential random. Various attempts to reveal the formation of enzyme-bound arginyl-adenylate provided no evidence for its existence. The reverse reaction, i.e., the deacylation of arginyl-tRNA, required both AMP and PP1. This observation is consistent with the mechanism proposed.抄録全体を表示PDF形式でダウンロード (722K) -
Yoshihide OHE, Hiroaki HAYASHI, Koichi IWAI1986 年 100 巻 2 号 p. 359-368
発行日: 1986年
公開日: 2008/11/18
ジャーナル フリーThe complete amino acid sequence of a main variant, Hlb, of human spleen histone Hl was determined, following previous determinations of human spleen histones H2B, H2A, H3, and H4. High-performance liquid chromatography on C8 silica of the H1 fraction yielded the homogeneous Hlb subfraction; this variant was estimated to account for 60% of the total of the four H1 variants. The sequence determination was performed with four main fragments, I to IV, obtained by limited chymotryptic digestion of H1b. Together with direct sequencing by automated Edman degradation of fragments II, III, and IV, fragment I, blocked at the N-terminal, and fragment IV, the C-terminal half the Hlb molecule, were sequenced after further digestion with staphylococcal protease and others. The four fragments were aligned with three overlapping peptides each derived from chymotryptic partial fragments, I-II and I-II-Tll, and intact H1b. Carboxypeptidase digestion of intact Hlb confirmed the C-terminal sequence of the molecule. Thus, the total sequence of Hlb was completely determined; it consists of a total of 218 amino acid residues, has a molecular weight of 21, 734 in the unmodified form, and is completely acetylated at the N-terminal serine residue and partially methylated at the lysine residue 25. This sequence is compared with two mammalian somatic HI sequences.抄録全体を表示PDF形式でダウンロード (609K) -
Hiroyuki KOMIYA, Masami HASEGAWA, Shosuke TAKEMURA1986 年 100 巻 2 号 p. 369-374
発行日: 1986年
公開日: 2008/11/18
ジャーナル フリーIn some amphibians and bony fishes, oocyte- and somatic-type 5S rRNA genes are expressed differently in oocytes and somatic cells. In order to determine at what stage of animal evolution this differential expression system appeared and how it is regulated, the sequences of oocyte and somatic 5S rRNAs from three invertebrates (sea urchin, sea hare, and silkworm) and two vertebrates (lamprey and chick) were analyzed. It was found that the oocyte 5S rRNA from lamprey consists of two components, while its somatic 5S rRNA consists of only one. In other animals, such differential expression of 5S rRNA in oocytes and somatic cells was not seen. A phylogenetic tree of 63 animal 5S rRNAs was constructed by means of the maximum parsimony method, and the evolution of oocyte and somatic-type 5S rRNAs was discussed.抄録全体を表示PDF形式でダウンロード (1063K) -
Structure and Properties of Albumin Tokushima and Its Proteolytic Processing by Cathepsin B In VitroYoshiko MATSUDA, Fumitaka OGUSHI, Koichi OGAWA, Nobuhiko KATUNUMA1986 年 100 巻 2 号 p. 375-379
発行日: 1986年
公開日: 2008/11/18
ジャーナル フリーAlbumin Tokushima is a Japanese genetic variant of human serum albumin. Two homozygous and 6 heterozygous subjects with this variant were found in a family. Albumin Tokushima was purified from sera of the homozygous subjects. Its amino acid composition and amino-terminal sequence were determined and compared with those of a normal serum albumin. Albumin Tokushima with the amino-terminal sequence of Arg-Gly-Val-Phe-His-Arg-Asp-Ala-His-Lys-Ser-Glu-Val-Ala-His-Arg-Phe-Lys-Asp-Leu-Gly-Glu-Glu-Asn-Phe was found to be the same abnormal pro-albumin as proalbumin Lille (Abdo, Y. et al. (1981) FEBS Lett. 131, 286-288). The isoelectric points of albumin Tokushima were pH 4.70 and 4.90 as compared with pH 5.05 and 5.25 of a normal serum albumin. Albumin Tokushima was converted to normal serum albumin by purified cathepsin B in vitro. Albumin Tokushima can bind Ni2+ at 4°C but binds little at 37°C.抄録全体を表示PDF形式でダウンロード (1264K) -
Isao KASUGAI, Michiyuki YAMADA1986 年 100 巻 2 号 p. 381-388
発行日: 1986年
公開日: 2008/11/18
ジャーナル フリーMyeloperoxidase synthesis during induction of differentiation of human promyelocytic leukemia HL-60 cells by 12-O-tetradecanoylphorbol-13-acetate (TPA) was studied. Differentiation was characterized by morphological changes, arrest of cell proliferation, development of cell adherence, and increased secretion of lysozyme. The cellular myeloperoxidase activity decreased early during induction of differentiation by TPA. Pulse-labeling experiments indicated that the rate of myeloperoxidase synthesis decreased to an undetectable level in cells exposed to TPA for 22h. The relative amounts of myeloperoxidase mRNA in TPA-treated and untreated cells were determined by measuring translatable mRNA activity in a reticulocyte lysate system. Reduction in the myeloperoxidase mRNA level was observed as early as after 3h treatment with TPA, and no myeloperoxidase mRNA was detected after 24h. Time course experiments indicated that the time required for 50% reduction of myeloperoxidase mRNA in TPA-treated cells was approximately 5h. These results suggest that TPA induces decrease of myeloperoxidase activity in HL-60 cells at a pretranslational level.抄録全体を表示PDF形式でダウンロード (1899K) -
Kiyoshi KAWAKAMI, Toshiko OHTA, Hiroshi NOJIMA, Kei NAGANO1986 年 100 巻 2 号 p. 389-397
発行日: 1986年
公開日: 2008/11/18
ジャーナル フリーClones carrying cDNA sequences for the α-subunit of the Na, K-ATPase from HeLa cells have been isolated. Nucleotide sequence analysis of the cloned cDNA has revealed the primary structure of this polypeptide, which consists of 1, 023 amino acids. The α-subunit of the human Na, K-ATPase exhibited 87% homology with its Torpedo counterpart and 98% homology with its sheep counterpart. The six putative transmembrane segments M1-M6 showed higher conservation than the total segments. Total genomic Southern hybridization indicated the existence of at most two copies, possibly only one, of the gene encoding the Na, K-ATPase α-subunit in the human genome.抄録全体を表示PDF形式でダウンロード (2184K) -
Nobuo MAKINO1986 年 100 巻 2 号 p. 399-405
発行日: 1986年
公開日: 2008/11/18
ジャーナル フリーAs an approach to elucidate the mechanism of the protein structure change in the cooperative ligand binding, the UV difference and CD spectra of aromatic residues in Panulirus iaponicus (spiny lobster) hemocyanin were examined.
The native hemocyanin showed an O2-induced narrow-banded change in the absorption spectrum around 290nm, which was not affected by pH in the range of 7.5 to 9.5. When the native hexameric protein was stripped of divalent cations with EDTA (at pH 7.5), the magnitude of the narrow-banded difference was reduced to about half, whereas it was almost completely abolished on dissociation into subunits (stripped at pH 9.5).
The magnitude of the absorption change was found to be proportional to the degree of O2 saturation in the native and stripped hemocyanins. It was inferred that the spectral difference reflects a tertiary structure change directly linked to the oxygenation, though it depends greatly on the subunit association.
Panulirus hemocyanin showed negative CD bands in the region of 260 to 300 nm, the intensities of which were considerably reduced by oxygenation and also by dissociation into subunits.抄録全体を表示PDF形式でダウンロード (508K) -
Iwao OHISHI1986 年 100 巻 2 号 p. 407-413
発行日: 1986年
公開日: 2008/11/18
ジャーナル フリーC2 toxin (C2T) elaborated by Clostridium botulinum types C and D is composed of two separate protein components, designated components I and II, which individually have little activity, but, when mixed and treated with trypsin, exert the potent activity. The present study provides the evidence that component I of the toxin catalyzes the hydrolysis of NAD into nicotinamide and ADP-ribose, whereas component II does not, indicating that component I of C2T has NAD-glycohydrolase activity, which ability is shared with cholera and diphtheria toxins. However, C2T affected neither glycerol production of fat cells nor protein synthesis in cell-free system. Component I of C2T in the presence of [α-32P]NAD radiolabeled a protein of Mr 46, 000 in the supernatant fractions of mouse tissue homogenates; the protein was abundant in brain, lung and intestine, whereas there was little or none of the protein in muscle. These results indicate that component I can catalyze the covalent attachment of the ADP-ribose moiety of NAD to intracellular protein, which differs from those modified with cholera and diphtheria toxins. The present data, together with previous findings, suggest that the biological activity of C2T is elicited by ADP-ribosylation activity of component I, which is internalized into the cells after binding to the receptor site introduced with the binding of component II to the cell surface membrane.抄録全体を表示PDF形式でダウンロード (1388K) -
Atsushi HARA, Tamotsu TAKETOMI1986 年 100 巻 2 号 p. 415-423
発行日: 1986年
公開日: 2008/11/18
ジャーナル フリーPositive ion fast atom bombardment (FAB) mass spectrometry of galactopsychosine and glucopsychosine was capable of showing not only the pseudo molecular ion peaks, but also various fragment ion peaks such as protonated sphingosine and its fragment ions. The percent distribution of sphingosine and dihydrosphingosine in each lysosphingolipid was determined by GLC of the trimethyl-silylated derivatives of long chain bases after methanolysis and was comparable to the relative intensities of ion peaks derived from the sphingosine and dihydrosphingosine groups. The FAB mass spectra showed that during the fast atom bombardment the sphingosine more preferentially gave rise to one and/or two fragment ions by loss of one and/or two molecules of water than the dihydrosphingosine did. The stereoisomerism of sphingosylphosphorylcholine containing mainly L-threo-sphingosine could be reconfirmed by carbon-13 NMR spectroscopy. Furthermore, although the carbon-13 NMR signals of sphingosine C-1, C-2, C-3, C-4, and C-5 showed significant chemical shift differences between D-erythro and L-threo-sphingosines of lysosphingolipids, it was concluded that the signal position of sphingosine C-3 was the most important for the determination of D-erythro and L-threo configuration in the long chain base moieties of lysosphingolipids.抄録全体を表示PDF形式でダウンロード (648K) -
Takashi IMAI, Takeshi CHO, Hiroyuki TAKAMATSU, Hitoshi HORI, Michiya S ...1986 年 100 巻 2 号 p. 425-432
発行日: 1986年
公開日: 2008/11/18
ジャーナル フリーDNA sequences encoding Ile-Glu-Gly-Arg and human prorenin were joined and placed under the transcriptional control of the Escherichia coli trp promoter-operator in the expression vector pTR501. E. coli cells transformed with pTR501 expressed high levels (30%. of total cell protein) of prorenin as part of a hybrid protein with the trp E gene product. The chimeric protein, accumulated in a sedimentable form, was dissolved in 6M guanidine HCI, purified to near homogeneity, and renatured by dialysis. The complete prorenin sequence was then excised from the renatured hybrid protein using blood coagulation factor Xa, a proteinase which is highly specific for the tetrapeptide insert Ile-Glu-Gly-Arg introduced between the 9 aminoterminal residues of the trp E gene product and the first amino acid (Thr 1) of prorenin. Human prorenin thus obtained was readily activatable with trypsin and showed close similarities to naturally occurring prorenin in its biochemical and immunochemical properties.抄録全体を表示PDF形式でダウンロード (2084K) -
Yoshio MISUMI, Kimirnitsu ODA, Yukio IKEHARA1986 年 100 巻 2 号 p. 433-438
発行日: 1986年
公開日: 2008/11/18
ジャーナル フリーWe have examined the effect of colchicine on the induction of alkaline phosphatase and its transport to the cell surface in a primary culture of rat hepatocytes. When freshly isolated hepatocytes were subjected to primary culture, alkaline phosphatase activity increased linearly starting at 6h and reached a maximum level (about 10 times the initial activity) at 24h after seeding. Radioimmunoassay with 125I-(anti-alkaline phosphatase)-IgG confirmed that the increase in enzyme activity was due to the increased amount of enzyme protein. The presence of colchicine in the culture medium (10-50μm) did not cause an additive effect on the enzyme induction, in contrast to the previous results obtained in in vivo experiments (Ikehara, Y. et al. (1978) J. Biochem. 84, 1335-1338; Oda, K. & Ikehara, Y. (1981) Biochim. Biophvs. Acta 640, 398-408). However, translocation of the induced enzyme to the cell surface was inhibited by colchicine in a dose-dependent manner. These results suggest that the enzyme induction by colchicine observed in vivo might not be due to its direct effect on hepatocytes, and that microtubules are involved in intracellular transport of the newly synthesized membrane protein.抄録全体を表示PDF形式でダウンロード (1204K) -
Yoshiharu ITOH, Sumiko KIMURA, Tsuneo SUZUKI, Kazuyo OHASHI, Koscak MA ...1986 年 100 巻 2 号 p. 439-447
発行日: 1986年
公開日: 2008/11/18
ジャーナル フリーNative connectin was isolated from porcine cardiac muscle using the method developed for the preparation of native connectin from chicken breast muscle (Kimura et al. (1984) J. Biochem. 96, 1947-1950). It was not necessary to keep cardiac muscle at 0°C before preparation: the proteolysis of α-connectin to β-connectin proceeded during the preparation of myofibrils. Cardiac connectin showed almost the same properties as those of skeletal muscle connectin: mobility in SDS gel electrophoresis, filamentous structure under an electron microscope, circular dichroism spectra, UV absorption spectra, and amino acid composition. Porcine cardiac connectin cross-reacted with antiserum against chicken breast muscle connectin as revealed by an immunoblot method. Immunoelectron microscopical observations revealed an abundance of connectin antigenic sites around the A-I junction area of cardiac myofibrils. Cardiac connectin also interacted with myosin and actin fila-ments at low ionic strengths to form aggregates. The extent of interaction was somewhat weaker in the case of cardiac connectin than skeletal muscle connectin, regardless of the origin of myosin and actin (porcine cardiac and rabbit skeletal muscles). In conclusion, cardiac connectin is very similar, but not identical to skeletal muscle connectin.抄録全体を表示PDF形式でダウンロード (4113K) -
Ikuko SAGAMI, Tetsuo OHMACHI, Hiroshi FUJII, Minro WATANABE1986 年 100 巻 2 号 p. 449-457
発行日: 1986年
公開日: 2008/11/18
ジャーナル フリーA major form of pulmonary cytochrome P-450 (pulmonary P-450MC) was purified approximately 165-fold from lung microsomes of 3-methylcholanthrene (MC)-treated hamsters. The purified preparation contained 14.2nmol of cytochrome P-450 (P-450) per mg protein and was essentially free from NADPH-cytochrome P-450 (cytochrome c)-reductase (NADPH-reductase) and epoxide hydrolase. Pul-monary P-450MC exhibits an absorption maximum at 446.5 nm in the difference spectrum of reduced hemoprotein-CO complex, and a low-spin state of ferric iron in the heme. By sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis, the molecular weight of pulmonary P-450MC was estimated to be 56, 000. In a reconstituted system, pulmonary P-450mc efficiently catalyzed benzo(a)pyrene (BP) hydroxylation, but showed low activities for 7-ethoxycoumarin O-deethylation and benzphetamine N-demethylation. In Ouchterlony double diffusion analysis, hamster pulmonary P-450MC reacted to the antibody prepared against rat hepatic P-450MC to form a faint precipitation line with a spur, indicating that the two P-450MCS have a common antigenic site but are not immunologically identical. When incubated with [14C]BP in a reconstituted system containing NADPH-reductase and epoxide by-drolase, hamster pulmonary P-450MC formed much higher amounts of BP diols, especially 7, 8-diol, than were formed by rat pulmonary P-450MC.抄録全体を表示PDF形式でダウンロード (2183K) -
Tamotsu HOMMA, Motoo WATANABE, Sachio HIROSE, Akira KANAI, Kenji KANGA ...1986 年 100 巻 2 号 p. 459-467
発行日: 1986年
公開日: 2008/11/18
ジャーナル フリーA radioimmunoassay for chicken calcitonin in chicken ultimobranchial glands was established utilizing a rabbit antiserum against eel calcitonin. This assay method, which is about 100 times as sensitive as the usual bioassay for hypocalcemic activity, was used for monitoring chicken calcitonin during its purification. The immuno-reactivity in chicken ultimobranchial extract was separated by SP-Sephadex C-25 chromatography into two fractions. Chicken calcitonin I, which was occurred in the major immunoreactive fraction, was further purified to homogeneity as shown by reverse phase HPLC. In the end, 39 nmol of chicken calcitonin I was obtained from 3, 384 chickens following a 12, 000-fold purification. The complete amino acid sequence of purified chicken calcitonin I was determined to be H-Cys-Ala-Ser-Leu-Ser-Thr-Cys-Val-Leu-Gly-Lys-Leu-Ser-Gln-Glu-Leu-His-Lys-Leu-Gln-Thr-Tyr-Pro-Arg-Thr-Asp-Val-Gly-Ala-Gly-Thr-Pro-NH2 and confirmed by synthesis. The specific biological activity of chicken calcitonin I (4, 500 MRCU/mg) was identical to that of eel calcitonin, which has the highest specific biological activity among the calcitonins so far isolated. Chicken calcitonin I resembled the calcitonins from the ultimobranchial glands both of salmon and eel in sequence, biological activity, and immunological property.抄録全体を表示PDF形式でダウンロード (695K) -
Chege J. MUKURIA, Yoichi FUJII, Shiro KATO, Masaharu NAIKI1986 年 100 巻 2 号 p. 469-475
発行日: 1986年
公開日: 2008/11/18
ジャーナル フリーThe specificities of five heterophile Hanganutziu and Deicher (HD) antibody-con-taining sera from four different cancer patients and one other diseased patients were compared. Three glycosphingolipids and one glycoprotein antigens and their chemi-cally modified derivatives were used.
1. The antibodies of all whole sera showed similar specificities.
2. IgG and IgM antibody fractions of each serum were separated. Although antibodies of the same class showed similar specificities, differences were detected between the specificities of IgG and IgM. IgG antibody specificities were dependent on the hydrophobic (ceramide) group while IgM antibodies were directed more to the terminal sialic acid moiety of the glycosphingolipid antigens.
3. The results suggested that a similar population of IgG-producing lympho-cytes is stimulated in patients.
Due to the similarities in specificities of HD antibodies, the results of this study will facilitate the future isolation of either IgG or IgM antibody-producing lym-phocyte(s) from a patient with HD antibodies and the establishment of a mono-clonal antibody through hybridization with a human rnyeloma cell line.抄録全体を表示PDF形式でダウンロード (519K) -
Hirofumi KUWABARA, Ivan VIDEN, Masahiko TSUCHIYA, Junko SUGATANI, Kuni ...1986 年 100 巻 2 号 p. 477-484
発行日: 1986年
公開日: 2008/11/18
ジャーナル フリーSeveral phosphatidyicholines (PC) and a phosphatidylethanolamine (PE) were subjected to liquid ionization (LI) mass spectrometry, in which a sample is ionized through energy transfer from metastable argon atoms under atmospheric pressure. Commercially available and synthesized, saturated or unsaturated fatty acid con-taining phospholipids and their mixtures were studied. A sample either as a con-centrated chloroform-methanol solution or with glycerol (matrix) gave characteristic peaks such as MH+ and four fragment ions. One of the fragment ions (e.g., m/z 551 of PC 16:0, 16:0) containing both fatty acid residues has been commonly observed with other ionization methods such as CI, FD, and FAB, but the other fragment ions have not been observed in other mass spectra with one exception on desorption CI. Ions b and d (e.g., m/z 464 and 328, respectively, for PC 16:0, 16:0) contain one fatty acyl residue and the other ion containing the phosphoryl-choline moiety appears at m/z 196 for PC. Thus the masses of the MH+ ion and these fragment ions provide useful structural information even in the case of a mixture. The ion b (e.g., m/z 488 of PC 18:0, 18:2) observed during an early period of heating was formed mainly by the loss of one acyl group at sn-1 of the glycerol backbone and thus may be used to differentiate the positional specificities of the constituent fatty acids. The temperature of the sample, however, should be controlled precisely, because it has a significant effect on the mass spectrum. The present method (LI) also provided useful information for a mixture of PC and PE.抄録全体を表示PDF形式でダウンロード (611K) -
Shinkichi TANIGUCHI, Shigehiro MORIKAWA, Haruki HAYASHI, Kimio FUJII, ...1986 年 100 巻 2 号 p. 485-491
発行日: 1986年
公開日: 2008/11/18
ジャーナル フリーThe effects of Ca2+ on ethanolaminephosphotransferase [EC 2.7.8.1] and choline-phosphotransferase [EC 2.7.8.2] activities in rabbit platelet membranes were studied using endogenous diglyceride and CDP-[3H]ethanolamine or CDP-[14C]choline as substrates. Both transferases required Mn2+, Co2+, or Mg2+ as a metal cofactor and the optimal concentrations of the metals for both activities were about 5, 10, and 5mM, respectively. When 5mM Mg2+ was used as a cofactor, both transferase activities were inhibited by a low concentration of Ca2+ (half maximal inhibition at approx. 15μm). In the presence of 5mM Mn2+, however, approx. 5mM Ca2+ was required to produce half maximal inhibition. The Ca2+-induced inhibition was reversible and the rate of the inhibition was not affected either by the concentrations of the CDP-compound or by exogenously added diacylglycerol. The relationship between Ca2+ and both Mg2+ and Mn2+ on the transferase activities was competitive. 45Ca2+ binding (and/or uptake) to the platelet membranes was inhibited by Mn2+, Mg2+, and Co2+, in a concentration-dependent manner. However, the inhibitory effects of the three metal ions on the total Ca2+ binding (and/or uptake) did not correlate with the activation of both transferase activities by the three metal ions in the presence of Ca2+. These results suggest that both transferase activities are regulated by low concentrations of Ca2+ in the presence of optimal concentrations of Mg2+, and that the inhibition is mediated directly by Ca2+, which interacts with a specific metal cofactor binding site(s) of the transferases.抄録全体を表示PDF形式でダウンロード (574K) -
Kohji ITOH, Kohji UENO, Shunji NATORI1986 年 100 巻 2 号 p. 493-498
発行日: 1986年
公開日: 2008/11/18
ジャーナル フリーPreviously, we showed that 20-hydroxyecdysone induces selective phosphorylation of a fat body protein of Sarcophaga peregrina with a molecular mass of about 30, 000 (30-kDa protein) (Itoh, K., Ueno, K., & Natori, S. (1985) Biochem. J. 227, 683-688). This paper describes the identification of this 30-kDa protein. From the electro-phoretic profile of 40S ribosomal proteins on two-dimensional polyacrylamide gel electrophoresis, the 30-kDa protein was identified as S6.抄録全体を表示PDF形式でダウンロード (2081K) -
Kazuo HARADA, Mamoru TAMURA, Isao YAMAZAKI1986 年 100 巻 2 号 p. 499-504
発行日: 1986年
公開日: 2008/11/18
ジャーナル フリーUpon the addition of hydrogen peroxide or ethyl hydroperoxide to sperm whale metmyoglobin (MbIII) in the presence of ethanol, MbIII was converted to oxymyo-globin (MbIIO2) under aerobic conditions and carboxymyoglobin (MbIICO) under CO-saturated conditions. MbIIO2 and MbIICO were also formed when ethanol was added to ferryl myoglobin (MbIV) which had been formed from the reaction of MbIII with hydrogen peroxide. From the stoichiometry, the primary reaction was formulated as follows.
MbIV +ethanol→MbII+acetaldehyde
The reaction was optimal at pH 7.0-7.5. Sperm whale ferryl myoglobin was reduced less effectively by methanol and n-butanol, but not at all by sec- and tert-butanols. The reduction of ferryl hemoproteins by ethanol was slower with horse heart myoglobin and was not observed with bovine hemoglobin or horseradish peroxidase.抄録全体を表示PDF形式でダウンロード (439K) -
Yasunori KOZUTSUMI, Toshisuke KAWASAKI, Ikuo YAMASHINA1986 年 100 巻 2 号 p. 505-512
発行日: 1986年
公開日: 2008/11/18
ジャーナル フリーOligosaccharides of a non-oligomannoside type were released from porcine α-mannosidase by hydrazinolysis, and were fractionated into at least 15 homogeneous oligosaccharides. Most of them are oligosaccharides with galactose and N-acetyl-glucosamine residues attached to a common core, αMan2βManβGlcNAc(±α-L-Fuc)βGlcNAc. About 50% of the oligosaccharides contain one or two outer chains composed of one β-linked N-acetylglucosamine and two β-linked galactose residues attached to the core portions, and the others seem to be metabolic inter-mediates. Based on the results of studies on the binding of α-mannosidase to RCA (Ricinus communis agglutinin) I-agarose and MBP (mannan-binding protein)-Se-pharose, which are specific for glycoproteins possessing N-acetyllactosamine-type and oligomannoside-type (including oligomannosides with N-acetylglucosamine at the reducing termini) oligosaccharides, respectively, about 85% of the enzyme molecules were found to have both types of oligosaccharides. Similarly, it was shown that of the several acid hydrolases present in the lysosomes purified from rat liver, only α-mannosidase has both types of oligosaccharides, and the greater parts of β-glucuronidase, acid phosphatase and β-N-acetylhexosaminidase seem to have only oligomannoside-type oligosaccharides.抄録全体を表示PDF形式でダウンロード (617K) -
Bunzo MIKAMI, Keiichi NOMURA, Yuhei MORITA1986 年 100 巻 2 号 p. 513-516
発行日: 1986年
公開日: 2008/11/18
ジャーナル フリーThe blocked N-terminus and N-terminal sequence of soybean β-amylase were determined by analyzing the acidic peptides derived on peptic digestion of the enzyme. The acidic peptides were separated from the digest on a Dowex 50×2 column and purified by reversed phase-high performance liquid chromatography (RP-HPLC). The major acidic peptide, Pep-4, was a heptapeptide with a molecular weight of 766. Forty-eight hundredths mol acetyl group and 0.61mol acetyl-Ala per mol of Pep-4 were detected on RP-HPLC analysis. The N-terminal 9 amino acid sequence of soybean β-amylase was deduced to be acetyl-Ala-Thr-Ser-Asp-Ser-Asn-Met-(Gly-Leu) from the results of sequence analysis of Pep-4 and amino acid analysis of other acidic peptides.抄録全体を表示PDF形式でダウンロード (296K) -
Yoshimichi ONOYAMA, Iwao OHTSUKI1986 年 100 巻 2 号 p. 517-519
発行日: 1986年
公開日: 2008/11/18
ジャーナル フリーOne of the chymotryptic troponin T subfragments, troponin T2, reconstituted approximately 80% of the Ca2+-sensitizing action of troponin T on actomyosin in the presence of troponin C, troponin I, and tropomyosin. This indicates that the Ca2+-regulatory property of troponin T is mostly localized in the troponin T2 region of the C-terminal 101 residues.抄録全体を表示PDF形式でダウンロード (190K)
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