巻号一覧
106 巻, 4 号
選択された号の論文の35件中1～35を表示しています
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Toshimi Tamaki, Sueharu Horinouchi, Masahiro Fukaya, Hajime Okumura, Y ...1989 年 106 巻 4 号 p. 541-544
発行日: 1989年
公開日: 2011/01/25
ジャーナル フリーThe nucleotide sequence of the membrane-bound aldehyde dehydrogenase (ALDH) gene from an industrial vinegar producer, Acetobacter polyoxogenes, was determined. Comparison of the sequence with the NH2-terminal amino acid sequence of the mature ALDH and determination of the actual translational initiation codon by means of in vitro manipulation of the upstream and proximal regions of the cloned gene showed that ALDH was primarily translated as a 773-amino-acid protein and that the 44-amino-acid sequence at the NH2-terminus, which probably serves as a signal peptide, was processed during maturation and localization in the membrane. When ALDH was expressed in a large quantity in Escherichia coli cells after the coding region had been placed downstream of the lac promoter, the ALDH protein, which still contained the signal peptide and had no ALDH activity, was localized in the membrane fraction.抄録全体を表示PDF形式でダウンロード (1510K) -
Masayuki Komada, Ichiro Kudo, Hiroshi Mizushima, Naomi Kitamura, Keizo ...1989 年 106 巻 4 号 p. 545-547
発行日: 1989年
公開日: 2011/01/25
ジャーナル フリーThree cDNA clones coding for rat platelet phospholipase A2 and a homologous protein were isolated from a rat megakaryocyte cDNA library and sequenced. One (prPLA2-1) carries a 708 nucleotide long insert. The others (prPLA2-2 and -3) differ from clone 1 in three nucleotides and have a 748 nucleotide long insert. All contain a single open reading frame which encodes a 146 amino acid long polypeptide. Based on the deduced amino acid sequence, we concluded that prPLA2-1 encodes rat platelet phospholipase A2. prPLA2-2 and -3 most probably encode a protein homologous to phospholipase A2 with two amino acid replacements. A typical signal peptide sequence (21 amino acid long), located at the NH2 termini of the deduced structure, was immediately followed by a polypeptide which corresponds to the mature enzyme, suggesting that the rat platelet enzyme is not expressed as a proenzyme form. Northern blot analysis showed a single transcript, which is 900 to 1, 100 nucleotides long, in the poly (A) +RNA fractions of rat megakaryocytes, bone marrow cells, peritoneal cells of caseinate-treated rats, and spleen cells.抄録全体を表示PDF形式でダウンロード (1328K) -
Masanori Mitta, Kiyozo Asada, Yuka Uchimura, Fusao Kimizuka, Ikunoshin ...1989 年 106 巻 4 号 p. 548-551
発行日: 1989年
公開日: 2011/01/25
ジャーナル フリーTwo overlapping cloned cDNAs encoding the entire amino acid sequences of the subunits of acylamino acid-releasing enzyme (AARE)[EC 3.4.19.1] have been isolated from porcine liver cDNA λgt10 cDNA libraries and sequenced. Sequence analyses of the cDNA and several Achromobacter protease I-digested peptides of the purified protein revealed that porcine liver AARE consists of four identical subunits, and each comprising a single chain of 732 amino acids with acetylmethionine at the N-terminus.抄録全体を表示PDF形式でダウンロード (752K) -
Akira Yamamoto, Hiroko Toda, Fumio Sakiyama1989 年 106 巻 4 号 p. 552-554
発行日: 1989年
公開日: 2011/01/25
ジャーナル フリーThe “vapor-phase” hydrazinolysis method was devised for the microdetermination of the carboxyl-terminal residue of a protein. With this method, a polypeptide sample is degraded with vaporized hydrazine. The optimum conditions for hen egg-white lysozyme were established to be 2 to 4 h at 90 or 100°C, the recovery of the carboxyl-terminal leucine being about 70%. With this vapor-phase method, side reactions are reduced and the time of hydrazinolysis is shortened. The limit of quantitation for the carboxyl-terminus of a protein is about 50pmol, as judged so far with hen egg-white lysozyme. The carboxyltermini of several proteins were determined using this novel procedure.抄録全体を表示PDF形式でダウンロード (1241K) -
Yuji Sugita, Takashi Tobe, Eiichi Oda, Motowo Tomita, Ko Yasukawa, Nob ...1989 年 106 巻 4 号 p. 555-557
発行日: 1989年
公開日: 2011/01/25
ジャーナル フリーHuman erythrocytes contain a membrane protein, MACIF, which inhibits the formation of a membrane attack complex (MAC) of complement. We have cloned and sequenced the complementary DNA of MACIF messenger RNA. The amino acid sequence predicted from its nucleotide sequence consists of 128 amino acids. The amino-terminal 25 residues may correspond to a signal peptide. The carboxy-terminal sequence confirmed that MACIF is a glycosylphosphatidylinositol (GPI)-anchored protein. The amino acid sequence of MACIF was partially determined by established techniques for protein chemistry and the resultant sequence was consistent with that predicted from the nucleotide sequence. The results of sequence analyses also suggested that asparagine at the 18th position was N-glycosylated. When mRNA obtained from the MACIF cDNA clone with SP6 RNA polymerase was microinjected into Xenopus oocytes, the oocytes synthesized a product which exhibited MACIF activity and reacted with anti-MACIF antibody. Comparison of the predicted sequence revealed significant homology with mouse Ly-6 antigens.抄録全体を表示PDF形式でダウンロード (717K) -
So Iwata, Takashi Minowa, Bunzo Mikami, Yuhei Morita, Takahisa Ohta1989 年 106 巻 4 号 p. 558-559
発行日: 1989年
公開日: 2011/01/25
ジャーナル フリーL-Lactate dehydrogenase from Bifidobacterium longum aM101-2 was overexpressed in Escherichia coli and then purified. The enzyme was crystallized from a polyethylene glycol 6000 solution by the hanging drop vapor diffusion method. Crystals grown in the presence of NADH (type II), both NADH and oxamate (type III), and NADH, oxamate, and FBP (type IV) were analyzed. All three crystal forms belong to the orthorhombic system, space group P21212. The cell dimensions of the type II crystals were a=106.2Å, b=131.6Å, and c=63.8Å. Those of the type III and type IV crystals were a=106.4Å, b=131.4Å, and c=63.8 Å. The type III crystals diffract X-rays to beyond 2.5Å spacing. The type II and type III crystals were stable as to X-ray irradiation.抄録全体を表示PDF形式でダウンロード (930K) -
Yukako Hayashi, Reiko Urade, Shigeru Utsumi, Makoto Kito1989 年 106 巻 4 号 p. 560-563
発行日: 1989年
公開日: 2011/01/25
ジャーナル フリーThe cytoplasmic peptide elongation factor, EF-1α, is anchored at the endoplasmic reticulum membrane by phosphatidylinositol via ethanolamine bridging presumably to Asp306of the protein.抄録全体を表示PDF形式でダウンロード (1803K) -
Kosuke Abe, Kazuhiko Yamamoto, Hyogo Sinohara1989 年 106 巻 4 号 p. 564-568
発行日: 1989年
公開日: 2011/01/25
ジャーナル フリーThe interactions of mouse murinoglobulin and α-macroglobulin with several proteinases were investigated by gel filtration and by assays of amidolytic activity towards synthetic substrates in the presence of proteinaceous enzyme inhibitors as well as assays of the inhibition of proteolytic activity. Mouse α-macroglobulin formed complexes with thrombin, clotting factor Xa, plasmin, pancreatic kallikrein, plasma kallikrein, submaxillary gland trypsin-like proteinase, neutrophil elastase, and pancreatic elastase. These complexes lost the proteolytic activities against high-molecular-weight substrates, but protected the active sites of the enzymes from inactivation by their proteinaceous inhibitors. Mouse murinoglobulin showed essentially the same properties except (i) that it did not form a complex with the clotting factor Xa, and (ii) that it did not protect plasma kallikrein, neutrophil elastase or submaxillary proteinase from inactivation by their proteinaceous inhibitors, although it formed complexs with these proteinases. No interaction was detected between Clostridium histolyticum collagenase and murinoglobulin or α-macroglobulin. These results indicate (i) that murinoglobulin has a proteinase-binding spectrum similar to that of α-macroglobulin, but is weaker in the ability to protect the bound proteinases from inactivation by the proteinaceous inhibitors than α-macroglobulin and (ii) that mouse α-macroglobulin has essentially the same inhibitory spectrum as the human homologue.抄録全体を表示PDF形式でダウンロード (1307K) -
Tomohide Uno, Yoshio Imai1989 年 106 巻 4 号 p. 569-574
発行日: 1989年
公開日: 2011/01/25
ジャーナル フリーThe nucleotide sequence of cDNA for rabbit liver cytochrome P-450 (laurate (ω-1)-hydroxylase) was replaced with that for rabbit liver cytochrome P-450 (testosterone 16α-hydroxylase) in various regions coding for the amino acid sequence between residues 43 and 261. Six chimeric cDNAs thus constructed were cloned into expression vector pAAH5, and expressed in Saccharomyces cerevisiae AH22 cells under the control of yeast ADH1 promoter. Chimeric P-450s synthesized in the transformed yeast cells were purified partially and their catalytic and spectral properties were examined and compared with those of the chimeric P-450 which is considered to possess the same catalytic properties as the wild-type P-450. In the oxidized state the chimeric P-450s exhibited a low- and high-spin mixed-type absorption spectrum of cytochrome P-450 and the spectrum was converted to a typical high-spin type on addition of laurate or caprate, indicating the binding-of the fatty acids to the substrate site of the chimeric P-450s. However, the affinities of the fatty acids for the chimeras devoid of the sequence of P-450 (laurate (ω-1)-hydroxylase) in either of the regions spanning residues 90-125 and 210-261 were 10 to 20 times lower than those for the chimeras containing the sequence of the wild-type P-450 in both regions. The latter chimeras have about the same affinities as the chimera which is essentially the wild-type P-450. In the reconstituted system containing purified enzymes, the chimeras containing the sequence of the wild-type P-450 in both regions catalyzed (ω-1)-hydroxylation of the fatty acids, but their activities were 51-67% of that of the chimera which is essentially the wild-type P-450. The chimera which contains the sequence of the wild-type P-450 in the region covering residues 90-125 but not residues 210-261 was about one-tenth as active as the chimera which is essentially the wild-type P-450. On the other hand, the chimeras devoid of the sequence of the wild-type P-450 in the region spanning residues 90-125 were completely devoid of the hydroxylase activities. These results indicate that two segments of P-450 (laurate (ω-1)-hydroxylase) covering residues 90-125 and 210-262 constitute the substrate binding sites and cooperate to fix the fatty acid at an appropriate position on the P-450 molecule, and the former segment is essential to the hydroxylase activity. The reduced CO complex of the chimeric P-450s showed the typical absorption spectrum of cytochrome P-450. However, the CO complexes of the chimeras devoid of the sequence of the wild-type P-450 covering residues 90-125 were much more unstable to denaturation than those containing this sequence, suggesting that this sequence protects the conformation of the chimeric P-450s in the reduced state against denaturation.抄録全体を表示PDF形式でダウンロード (1817K) -
Isolation of Phage Clones Which Cover a Functionally Active Gene and Structural Analysis of the Region Upstream from the Translation Initiation CodonKeiichi Takeishi, Sumiko Kaneda, Dai Ayusawa, Kimiko Shimizu, Osamu Go ...1989 年 106 巻 4 号 p. 575-583
発行日: 1989年
公開日: 2011/01/25
ジャーナル フリーTwo genomic DNA fragments partially encoding human thymidylate synthase (TS)[EC 2.1.1.45] were previously cloned in λ phage from the mouse cell transformant, but had no transforming activity on mouse TS-negative mutant cells. In this study, an additional genomic DNA for human TS was cloned and demonstrated to have the transforming activity in combination with one of the two previously cloned DNAs and to produce human TS mRNA. The two transforming genomic DNAs overlapped and covered a region of 23kb in total. Using fragments from one of these DNAs, the structure of the 1.2-kb region around the ATG initiator codon of the TS gene was analyzed in relation to regulatory sequences of the gene. Sequence determination demonstrated the presence of an unusual inverted repeat consisting of a triple tandem repeat of a 28-bp sequence and an inverted sequence of the same length. These sequences can form three possible, stable, stem-loop structures, which may be interconvertible. Based on S1 nuclease mapping data and a line of circumstantial evidence, we deduced two major mRNA cap sites within the inverted sequence. Comparison of the human and mouse sequences upstream from the ATG initiator codon revealed many significant blocks of sequence homology, especially in the regions around the deduced cap sites.抄録全体を表示PDF形式でダウンロード (3521K) -
Takao Ohyashiki, Michihiro Taka, Tetsuro Mohri1989 年 106 巻 4 号 p. 584-588
発行日: 1989年
公開日: 2011/01/25
ジャーナル フリーThe effects of neuraminidase treatment on the membrane surface charge density and/or membrane potential of the porcine intestinal brush-border membrane vesicles were studied by using three fluorescent dyes, 1, 6-diphenyl-1, 3, 5-hexatriene (DPH), 1-anilino-8-naphthalene sulfonate (ANS), and 3, 3'-dipropyl-2, 2'-thiadicarbocyanine iodide (DiSC3 (5)). The results of quenching studies of DPH-labeled membranes using cationic T1+ and anionic (I-) quenchers suggested an increase of negative charge on the membrane surface by desialylation upon neuraminidase treatment. This interpretation was further supported by a decrease of ANS-binding affinity of the membranes after treatment with the enzyme. In addition, the degree of valinomycin-induced fluorescence change of DiS-C3 (5)-probed membranes in the presence of various concentrations of KCl was reduced by treatment of the membranes with neuraminidase. This suggests that penetration of the dyemolecules into the vesicle interior is facilitated by the treatment. The membrane potentials estimated from the null point of valinomycin-induced changes in the DiS-C3 (5) fluorescence of the control and neuraminidase-treated membranes were -25 to -29.7 and -40 to -48.8 mV respectively. From these results, it is suggested that the membrane surface charge density and/or membrane potential of the intestinal brush-border membranes are susceptible to modification of carbohydrate moieties on the membrane surface by neuraminidase treatment.抄録全体を表示PDF形式でダウンロード (1202K) -
Yuji Sugita, Toshio Mazda, Motowo Tomita1989 年 106 巻 4 号 p. 589-592
発行日: 1989年
公開日: 2011/01/25
ジャーナル フリーThe protein corresponding to P-18 (Sugita et al.(1988) J. Biochem. 104, 633-637) was isolated from native human erythrocyte, and newly designated membrane attack complexinhibitory factor (MACIF). The amino-terminal sequenceofthis protein was determined to be Leu-Gln-Cys-Tyr-Asn-Cys-Pro-Asn-Pro-Thr. Endoglycosidase F digestion of MACIF decreased its molecular weight by about 6K on SDS-PAGE. On the other hand, endoglycosidase H, neuraminidase, or endo-α-N-acetylgalactosaminidase treatment had no effect on the molecular weight, indicating that MACIF has complex-type N-linked oligosaccharide chains, but noO-linked chain. MACIF was highly resistant against trypsin digestion and heat treatment. The inhibitory activity of MACIF on the hemolysis of EC5-8 cells was comparable to that on EC5-7 cells, indicating that MACIF inhibited the binding of C9 to the intermediate cells, or the subsequent C9 polymerization.抄録全体を表示PDF形式でダウンロード (1501K) -
Keiko Tamiya-Koizumi, Hayato Umekawa, Shonen Yoshida, Kiyohide Kojima1989 年 106 巻 4 号 p. 593-598
発行日: 1989年
公開日: 2011/01/25
ジャーナル フリーA sphingomyelinase, which specifically hydrolyzes sphingomyelin into ceramide and phosphocholine, was solubilized from nuclear matrix fraction of rat ascites hepatoma, AH7974 cells. The solubilized enzyme was subjected to Mono Q column chromatography in an FPLC system. The sphingomyelinase which was adsorbed on the column and eluted at 0.25-0.5M NaCl was characterized. The enzyme required 10mM MgCl2, 0.01% Triton X-100, 1mM dithiothreitol, and a higher concentration of buffer than 1M for its maximal activity, and the optimal pH was 6.7-7.2 in 2M Tris/acetic acid or 7.5 in 2M potassium acetate/acetic acid. N-Ethylmaleimide completely inhibited the enzyme activity at 0.2mM. Therefore, this enzyme is classified as a Mg2+ -dependent, neutral sphingomyelinase. The sphingomyelinase sedimented at 4.3S through a 10-30% glycerol gradient containing 2M potassium acetate. This enzyme was highly specific to sphingomyelin and did not hydrolyze phosphatidylcholine, phosphatidylethanolamine, phosphatidylserine, and phosphatidylinositol. Various characteristics of the nuclear sphingomyelinase were similar to those of the plasma membrane enzyme except its requirement for a high concentrationofbuffer and SH-reagent.抄録全体を表示PDF形式でダウンロード (1109K) -
Tarou Ogurusu, Shigeo Wakabayashi, Takahide Watanabe, Munekazu Shigeka ...1989 年 106 巻 4 号 p. 599-605
発行日: 1989年
公開日: 2011/01/25
ジャーナル フリーWe investigated the reaction mechanism for GTP-dependent Ca2+ uptake by canine cardiac microsomes enriched in fragmented sarcoplasmic reticulum (SR), because previous studies reported that GTP utilization in cardiac SR occurs via a pathway very different from that for ATP utilization (for a review, see “Entman, M. L., Bick, R., Chu, A., Van Winkle, W. B., & Tate, C. A.(1986) J. Mol. Cell. Cardiol. 18, 781-792”). In cardiac microsomes, we detected slow but distinct oxalate-dependent Ca2+ accumulation, which reached 550nmol/mg protein in 10min, and similarly slow Ca2+ -dependent GTP hydrolysis. In 50μM [γ-32P]- GTP at 0°C, we detected Ca2+ -dependent formation of phosphoprotein whose level in the steady state was about ahalfofthe maximum obtained with [γ-32P] ATP. Kinetic properties of the phosphoprotein, its molecular weight and its chemical stabilityafter the acid treatment are consistent with the conclusion that the phosphoprotein is an acylphosphate intermediate for Ca2+ -dependent GTP hydrolysis catalyzed by the Ca2+ -pump ATPase. Analysis of the kinetics of the turnover of phosphoprotein revealed that slow GTP hydrolysis is due to slow phosphoprotein formaton; at 25°C, the latter arises mainly from slow binding of Ca2+ to the dephosphorylated enzyme. These results indicate that, contrary to the previous data, the reaction pathway for GTP-dependent Ca2+ transport in cardiac SR is basically the same as that for ATP-dependent transport.抄録全体を表示PDF形式でダウンロード (1304K) -
Kimiko Saeki, Makio Tokunaga, Hian Ann Ting, Takeyuki Wakabayashi1989 年 106 巻 4 号 p. 606-611
発行日: 1989年
公開日: 2011/01/25
ジャーナル フリーWe have developed a new method to prepare single-headed heavy meromyosin with high purity and a high yield. To examine whether the two heads on the same myosin molecule work cooperatively or not, it is important to prepare pure single-headed heavy meromyosin. Myosin was extracted from myofibrils treated with a solution containing CyDTA, a strong divalent cation chelator. CyDTA treatment was essential to the production of sHMM. Then such myosin was digested with chymotrypsin in the presence of divalent cations at high ionic strength. Crude sHMM was separated from double-headed HMM by affinity chromatography using an ADP-column. Contaminating Si was removed by gel filtration. Heavy chain of sHMM obtained by the present method had no nick. Purified sHMM showed normal EDTA-ATPase and Ca-ATPase. It interacted with thin filament and its ATPase was activated by actin normally.抄録全体を表示PDF形式でダウンロード (3731K) -
Masayuki Suda, Hiroaki Hayashi1989 年 106 巻 4 号 p. 612-615
発行日: 1989年
公開日: 2011/01/25
ジャーナル フリーTetrahymena pyriformis was starved in 50mM Tris-HCl, pH 7.5, at 28°C. The number of cells did not change appreciably under the starvation conditions. Nuclear proteins of unstarved cells and cells starved for 1, 2, 4, and 7 d were analyzed by SDS-polyacrylamide gel electrophoresis. Most of the large amount of nonhistone proteins present in the unstarved cell nucleus disappeared with the starvation time. However, the relative amounts of the high mobility group protein and histones did not change appreciably. On the other hand, a protein with a molecular weight of ca. 16, 000 gradually accumulated in the nucleus on starvation. This protein was extracted with 0.25M HCl, but was not soluble in 0.5M perchloric acid. The amino acid composition and molecular weight of this protein were similar to those of HMG protein LG-2 of T. thermophila. Some lysyl endopeptidase peptides of this protein were found to have amino acid sequences present in LG-2, thus we tentatively named it an LG-2-like protein.抄録全体を表示PDF形式でダウンロード (1878K) -
Manzoor Shah Ahmad, Rashid Ali1989 年 106 巻 4 号 p. 616-620
発行日: 1989年
公開日: 2011/01/25
ジャーナル フリーThis paper describes the antigenicity of pig kidney diamine oxidase [EC 1.4.3.6] and the possible role of constituent amino acids in the epitope structure of the enzyme. The loss of 62% of the biological activity after DAO-anti-DAO antibodies interaction was attributed to the steric hindrance caused by binding of antibody to the enzyme molecule. A gradual loss in antigenicity during ultraviolet (UV) irradiation was observed without any significant conformational change, demonstrating the destruction of antigenic determinants. However, ethoxyformylation of nine histidyl residues with complete inactivation caused no change in immunoreactivity. The results indicate that the antigenic sites and catalytic sites are located at different positions along the polypeptide chain. Moreover, the results of lysine residue modification were suggestive of possible involvement of lysine in the antigenic determinants of DAO.抄録全体を表示PDF形式でダウンロード (1680K) -
Yasumasa Gotoh, Yoshifusa Kondo, Hiroyuki Kaji, Atsushi Takeda, Tatsuy ...1989 年 106 巻 4 号 p. 621-626
発行日: 1989年
公開日: 2011/01/25
ジャーナル フリーBilirubin oxidase [EC 1.3.3.5], purified from the culture medium of Myrothecium verrucaria, was found to contain two blue copper atoms per protein molecule with a molecularweight of ca. 52 kDa. The two copper atoms were estimated to be in the all cupric state by the cuproine colorimetric method and also atomic absorption analysis. We could remove the reduced cuprous ions from the holo enzyme by adding ascorbate, followed by a KCN solution, yielding an apo-enzyme with no activity. The apo-enzyme can be reconstituted with Cu or other divalent cations such as Co, Fe, and Cd, with accompanying recovery of the enzyme activity. The activity recovery depended upon the species of cation employed; Cu being most effective, an almost 100% recovery, and Cd the least, only a 25% recovery. We could obtain information on the copper ions and their coordination structure by spectroscopic analyses of the apo- and reconstituted enzymes, obtaining such as absorption, CD, MCD, and XPS spectra. The bilirubin oxidase catalyzed-reaction was a second orderreaction with respect to copper bound with protein. The donor set was of the CuSSN2 (S=Cys, S=Met, N=His) type, i. e., the same as in the case of blue copper proteins. On studying the Co-substituted enzyme, it was revealed that the copper site of the enzyme had a 4-coordinated structure.抄録全体を表示PDF形式でダウンロード (1460K) -
Toshihiko Saheki, Motoyuki Shimonaka, Keiko Uchida, Takeshi Mizuno, Sh ...1989 年 106 巻 4 号 p. 627-632
発行日: 1989年
公開日: 2011/01/25
ジャーナル フリーThe presence of subtypes of atrial natriuretic peptide (ANP) receptor has been demonstrated by examining the immunological features and ligand specificities of the receptors in various bovine tissues. The antibody probe used was the antiserum raised against the bovine lung ANP receptor, and the tissues examined for the possible presence of different types of ANP receptor were the lung, kidney, adrenal cortex, ovary, choroid plexus, and vascular tissues. When incubated with Triton extracts of these tissues, the antiserum strongly cross-reacted with the ovary, kidney, and choroid plexus receptors as well as the homologous lung receptor (type I). The adrenal and vascular receptors were recognized only weakly, however, suggesting the presence of distinct ANP receptors (type II). In support of this immunochemical subtyping, type I and type II receptors showed a marked difference in their ability to bind the ANP analog atriopeptin I (ANP5-25): type I receptors in the lung exhibited a moderate affinity for atriopeptin I with a KD of 10-9M; however, type II receptors in adrenal and artery showed only a weak affinity for the analog with a KD of 10-6M. Structural analysis of affinity-labeled ANP receptors by SDS-PAGE indicated that type I and type II receptors have similar disulfide-linked dimeric structures. These results suggest that the integrated response of the circulatory system to ANP depends on at least two populations of the dimeric receptor: type I receptors play a major functionalrole in the kidney where ANP exerts diuretic and natriuretic effects, whereas type II receptors appear to have the predominant role in the adrenal and vascular beds where aldosterone secretion and vascular tone are modulated, respectively, by ANP.抄録全体を表示PDF形式でダウンロード (2434K) -
Makoto Nakagawa, Fumitake Tsukada, Toshihiro Nakayama, Kazuya Matsuura ...1989 年 106 巻 4 号 p. 633-638
発行日: 1989年
公開日: 2011/01/25
ジャーナル フリーDihydrodiol dehydrogenase activity was detected in the cytosol of various mouse tissues, among which kidney exhibited high specific activity comparable to the value for liver. The enzyme activity in the kidney cytosol was resolved into one major and three minor peaks by Q-Sepharose chromatography: one minor form cross-reacted immunologically with hepatic 3α-hydroxysteroid dehydrogenase and another with aldehyde reductase. The other minor form was partially purified and the major form was purified to homogeneity. These two forms, although different in their charges, were monomeric proteins with the same molecular weight of 39, 000 and had similar catalytic properties. They oxidized cis-benzene dihydrodiol and alicyclic alcohols as well as trans-dihydrodiols of benzene and naphthalene in the presence of NADP+ or NAD+, and reduced several xenobiotic aldehydes and ketones with NAD (P) H as a cofactor. The enzymes also catalyzed the oxidation of 3α- hydroxysteroids and epitestosterone, and the reduction of 3- and 17-ketosteroids, showing much lower Km values (10-7-10-6M) for the steroids than for the xenobiotic alcohols. The results of mixed substrate experiments, heat stability, and activity staining on polyacrylamide gel electrophoresis suggested that, in the two enzymes, both dihydrodiol dehydrogenase and 3 (17)α-hydroxysteroid dehydrogenase activities reside on a single enzyme protein. Thus, dihydrodiol dehydrogenase existed in four forms in mouse kidney cytosol, and the two forms distinct from the hepatic enzymes may be identical to3 (17)α-hydroxysteroid dehydrogenases.抄録全体を表示PDF形式でダウンロード (2071K) -
Kazuko Hori, Yoshihiro Yamamoto, Toshitaka Minetoki, Toshitsugu Kurots ...1989 年 106 巻 4 号 p. 639-645
発行日: 1989年
公開日: 2011/01/25
ジャーナル フリーThe entire gene for gramicidin S synthetase 1 (GS 1) was cloned into the plasmid vector p UC18, and the nucleotide sequences of the GS 1 gene and its flanking region were determined. The full-length clone was 4, 539 base pairs long and had an open reading frame of 3, 294 nucleotides coding for 1, 098 amino acids. The calculated molecular weight of 123, 474 agreed with the apparent molecular weight of 120, 000 found in SDS-PAGE of GS 1 from B. brevis. The nucleotide sequence of GS 1 gene was highly homologous to that of tyrocidine synthetase 1. The overall similarity between the deduced amino acid sequences of the two genes was 57.5%. The gene product of clone GS309 was easily purified to an essentially homogeneous state by ammonium sulfate fractionation followed by DEAE-Sepharose CL-6B, Ultrogel Ac A-34, and second DEAE-Sepharose CL-6B column chromatography. The purified protein catalyzed the D-phenylalanine-dependent ATP-32PP1 exchange reaction which is specific for GS 1 activity, and the specific activity of the purified product was nearly the same as the purified GS 1 from B. brevis. The product also showed a weak phenylalanine racemase activity.抄録全体を表示PDF形式でダウンロード (2676K) -
Kaoru Omichi, Kouichi Shiosaki, Kenichi Matsubara, Tokuji Ikenaka1989 年 106 巻 4 号 p. 646-650
発行日: 1989年
公開日: 2011/01/25
ジャーナル フリーThe mode of action of an α-amylase (yHXA) which was the gene product of a newly found human α-amylase gene expressed in yeast on synthetic substrates was compared with those of the gene products (yHSA and yHPA) of human salivary and pancreatic α-amylase gene in yeast. The substrates used were phenyl α-maltopentaoside (G5Φ) and its derivatives in which the CH2OH groups of the non-reducing-end glucose residues were converted to CH2NH2 (AG5Φ), COOH (CG5Φ), or CH2I (IG5Φ). The digests were subjected to HPLC to determine the amounts of products. The HPLC analysis revealed that yHXA and yHSA bound G5Φ to their active sites in similar manners to give the same products, while yHPA hydrolyzed it in a different way. Modifications of the non-reducing-end glucose of G5Φ caused change of the binding mode to the active sites of the enzymes. AGM, and CG5Φ were hydrolyzed by the enzymes to give more phenyl α-glucoside (GΦ) and less phenyl α-maltoside (G2Φ), while IG5Φ gave more G2Φ and less GΦ, compared with G5Φ. The substrate binding mode of yHXA changed more extensively than that of yHSA. The results suggested that there exists an amino acid replacement between yHXA and yHSA. The amino acid residues replaced are neither acidic nor basic, are located in subsite S3, and interact with the CH2OH residue of the non-reducing-end glucose residue of G5Φ. The residues appeared to be the 163rd in the enzyme amino acid sequence by reference to the data from the X-ray crystallographic studies of Taka-amylase A.抄録全体を表示PDF形式でダウンロード (1017K) -
Masao Miki1989 年 106 巻 4 号 p. 651-655
発行日: 1989年
公開日: 2011/01/25
ジャーナル フリーActin modified at Lys-61 with fluorescein 5-isothiocyanate (FITC) recovers the ability to polymerize following the binding of phalloidin. The resulting polymer (FITC-P-actin) activates the S1-Mg2+-ATPase activity to the same extent as non-labeled F-actin. However, in the absence of phalloidin, FITC-actin (0.5mg/ml) neither polmerized nor activated the S1-Mg2+-ATPase activity effectively even when it was preincubated with S1 for 3h in 0.1 m M ATP, 0.1m M CaCl2, and 1m M Tris/HCl (pH 8.0), in contrast to the previous report [Miller, L., Phillips, M., & Reisler, E.(1988) Eur. J. Biochem. 174, 23-29]. The modification of Lys-61 did not impair the ability to bind tropomyosin or tropomyosin-troponin. On the other hand, the fluorescence polarization of FITC-P-actin increased when tropomyosin or troponin-tropomyosin was added. Moreover, the modification of Lys-61 affected the regulation of the actin activation of the S1-Mg2+-ATPase activity by the tropomyosin and troponin complex. In 30m M KCl, 2.5m M ATP, and 5m M MgCl2, tropomyosin alone has been shown to inhibit the actin-activated Si-Mg2+-ATPase. This inhibition did not occur with FITC-P-actin even though tropomyosin was tightly bound. When troponintropomyosin was added, the FITC-P-actin activation of Si-Mg2+-ATPase activity was regulated in response to micromolar Ca2+ concentrations. On the othr hand, in 30m M KCl, 2.5m M ATP, and 2m M MgCl2, tropomyosin alone did not inhibit the actin-activated S1- Mg2+-ATPase activity with either non-labeled F-actin or FITC-actin. When troponintropomyosin was added, the ATPase activity was always inhibited to a certain extent with FITC-P-actin whether Ca2+ was present or not, although a strong inhibition in the absence of Ca2+ and no inhibition in the presence of Ca2+ were observed with non-labeled F-actin. These results indicate that Lys-61 is located in a region which is closely related to the regulation of the actin-myosin interaction by tropomyosin-troponin.抄録全体を表示PDF形式でダウンロード (1985K) -
Complete Primary Structures and Construction of Phylogenetic TreesKazuhiko Saeki, Yoshio Yao, Sadao Wakabayashi, Gwo-Jenn Shen, J. Grego ...1989 年 106 巻 4 号 p. 656-662
発行日: 1989年
公開日: 2011/01/25
ジャーナル フリーComplete amino acid sequences of ferredoxin and rubredoxin from Butyribacterium methylotrophicum, a methylotrophic hetero-acetogen, were determined by combination of protease digestion, Edman degradation, carboxypeptidase digestion, and/or partial acid hydrolysis. The ferredoxin was composed of 55 amino acids with a molecular weight of 5, 732 excluding iron and sulfur atoms and showed a typical 2 [4Fe-4S]-type ferredoxin sequence with an internal repeat at the 14-23 and 42-51 positions. The rubredoxin was composed of 53 amino acids with a molecular weight of 5, 672 excluding iron atom and showed a sequence similar to those of other anaerobic rubredoxins. The sequences were compared to those of corresponding proteins from six different bacteria to construct phylogenetic trees, which showed essentially the same topology. The relationships between the ferredoxin sequences from this bacterium and those of Clostridium thermoaceticum and Methanosarcina barkeri, both of which possess a carbonyl-dependent acetyl-CoA metabolic system, are also discussed.抄録全体を表示PDF形式でダウンロード (1596K) -
Toshiyuki Miyata, Fuminori Tokunaga, Takashi Yoneya, Katsuhiro Yoshika ...1989 年 106 巻 4 号 p. 663-668
発行日: 1989年
公開日: 2011/01/25
ジャーナル フリーTachyplesin is an antimicrobial peptide recently found in the acid extract of hemocytes from the Japanese horseshoe crab (Tachypleus tridentatus)[Nakamura, T. et al.(1988) J. Biol. Chem. 263, 16709-16713]. In our continuing studies on the peptide, we have found an isopeptide, tachyplesin II, and also polyphemusins I and II in hemocytes of the American horseshoe crab (Limulus polyphemus). The complete primary structures of these peptides, which are very similar to that of the previously isolated peptide, now named tachyplesin I, were determined to be as follows:
Polyphemusin I NH2-R-R-W-C-F-R-V-C-Y-R-G-F-C-Y-R-K-C-R-CONH2
Polyphemusin II NH2-R-R-W-C-F-R-V-C-Y-K-G-F-C-Y-R-K-C-R-CONH2
Tachyplesin II NH2-R-W-C-F-R-V-C-Y-R-G-I- C-Y-R-K-C-R-CONH2
The isopeptide, tachyplesin II, consists of 17 residues with a COOH-terminal arginine α-amide. On the other hand, both polyphemusins I and II were found to contain 18 residues due to an additional Arg residue at the NH, -terminal end as well as a COOH-terminal arginine α-amide. The disulfide linkages for polyphemusin I consisted of two bridges between Cys-4 and Cys-17 and between Cys-8 and Cys-13, which was identical to in the case of tachyplesin I. Moreover, all of these peptides inhibited the growth of not only Gramnegative and-positive bacteria but also fungi, such as Candida albicans M9. Furthermore, complex formation between these peptides and bacterial lipopolysaccharides was also observed in a double diffusion test. These results suggest that tachyplesins and polyphemusins are probably located in the hemocyte membrane, where they act on antimicrobial peptides as a self-defense mechanism in the horseshoe crab against invading microorganisms.抄録全体を表示PDF形式でダウンロード (1796K) -
Enzymic Characterization and Comparative Studies with Monoclonal AntibodiesKatsuyuki Imai, Takayuki Harada, Yasuo Takano, Shigeru Morikawa, Atsus ...1989 年 106 巻 4 号 p. 669-672
発行日: 1989年
公開日: 2011/01/25
ジャーナル フリーNeutral α-glucosidase was partially purified from granular fractions isolated from guinea pig polymorphonuclear leukocytes (PMNL). The native enzyme had a highmolecular weight, about 417, 000, with a subunit of 43, 000. The purified enzyme hydrolysed 4-methylumbelliferyl α-glucoside and maltose, but not isomaltose, trehalose, and glycogen. The enzyme was strongly inhibited by bromoconduritol and castanospermine, but only slightly by turanose. Monoclonal antibodies which can bind specifically to the enzyme were prepared by immunizing mice with the partially purified enzyme. Hybridomas producing the monoclonal antibodies were selected by an enzyme-linked immunosorbent assay. The seven monoclonal antibodies were found to react with the enzyme from PMNL, but not with the glycoprotein-processing α-glucosidase isolated from liver microsomes nor with the macrophage enzyme. The results indicated that PMNL contain a particulate neutral α-glucosidase enzymologically and imnunologically distinct from other α-glucosidases.抄録全体を表示PDF形式でダウンロード (1484K) -
Yasuhiko Konno, Shigeo Ohno, Yoshiko Akita, Hiroshi Kawasaki, Koichi S ...1989 年 106 巻 4 号 p. 673-678
発行日: 1989年
公開日: 2011/01/25
ジャーナル フリーA protein kinase C-related cDNA encodes a novel phorbol ester receptor/protein kinase, nPKCε, clearly distinct from the four “conventional” PKCs [Ohno, S., Akita, Y., Konno, Y., Imajoh, S., & Suzuki, K.(1988) Cell 53, 731-741]. We purified nPKCε from COS cells transfected with nPKC cDNA and compared its enzymatic properties with a conventional PKC, PKCα. nPKCε was eluted from a hydroxyapatite column at a position coincident with type II PKC and thus was separated from type III PKC (PKCα), the only PKC expressed in COS cells. The protein kinase activity of nPKCε is activated by phospholipids and diacylglycerols (or phorbol esters) in a manner similar to conventional PKCs. However, the cofactor dependencies and substrate specificites were clearly different from PKCα. A phospholipid, cardiolipin, enhances the kinase activity three- to fourfold compared with phosphatidylserine. The optimum Mg2+ concentration (3mM) is clearly different from those of conventional PKCs (10-20mM). The activation of nPKCε by these cofactors is totally independent of Ca2+. Similar to conventional PKCs, nPKCε autophosphorylates serine and threonine residues, indicating the specificity of the kinase to these amino acid residues. However, it shows a clearly different substrate specificity against exogenous substrates in that myelin basic proteins rather than histone are good substrates. These properties of nPKCε permit clear discrimination of nPKCε from conventional PKCs.抄録全体を表示PDF形式でダウンロード (2353K) -
Keishi Miwa, Michihiro Ohtsubo, Kimitoshi Denda, Toru Hisabori, Takaya ...1989 年 106 巻 4 号 p. 679-683
発行日: 1989年
公開日: 2011/01/25
ジャーナル フリーHomogeneous populations of hybrid α3β3γ complexes of the thermostable F1-ATPase containing one, two, or three copies of the mutationally impaired β subunits were produced using the solid phase reconstitution method. Two kinds of mutated β subunits were used for the reconstitution, one of which lacked the ability to bind any adenine nucleotides. The complexes containing one impaired β and two wild-type β subunits retained a significant amount of ATPase activity with cooperative kinetics, whereas those containing two or three impaired β subunits showed very little ATPase activity. These results imply that the catalysis of steady-state ATP hydrolysis can proceed even if one of the three β subunits in F1-ATPase is not functional.抄録全体を表示PDF形式でダウンロード (1139K) -
Kouichi Takimoto, Masato Okada, Hachiro Nakagawa1989 年 106 巻 4 号 p. 684-690
発行日: 1989年
公開日: 2011/01/25
ジャーナル フリーMembrane-bound inositolpolyphosphate 5-phosphatase was solubilized and highly purified from a microsomal fraction of rat liver. Its physicochemical and enzymological properties were compared with those of higyly purified preparations of two types of soluble enzyme (soluble Type I and Type II) from rat brain. The molecular masses of the membrane-bound and soluble Type I enzymes were 32kDa, while that of soluble Type II enzyme was 69kDa, as determined by molecular sieve chromatography. The membranebound and soluble Type I enzymes showed similar broad peaks on isoelectric focusing (pI 5.8-6.4), while soluble Type II enzyme showed multiple peaks in the region between pI 4.0-5.8. All three enzymes required divalent cation for activity. Mg2+ was the most effective for both the membrane-bound and soluble Type I enzymes, while Co2+ enhanced soluble Type II enzyme activity about 1.5-fold relative to Mg2+ at 1mM. The optimal pH of both the membrane-bound and soluble Type I enzymes was 7.8, while that of soluble Type II was 6.8. The Km values for inositol 1, 4, 5-trisphosphate [Ins (1, 4, 5) P3] of all three enzymes were similar (5-8μM), but those for inositol 1, 3, 4, 5-tetrakisphosphate [Ins (1, 3, 4, 5) P4] were quite different, the Km values of membrane-bound and soluble Type I enzymes being 0.8 μM, while that of soluble Type II was 130 μM. These similarities between the membranebound and soluble Type I enzymes suggest that these two molecules may be the same protein, and that concentrations of Ins (1, 4, 5) P3 and Ins (1, 3, 4, 5) P4, both of which are considered to play critical roles in the regulation of intracellular Ca2+-concentration, may be differently regulated by two functionally distinct enzymes.抄録全体を表示PDF形式でダウンロード (1825K) -
Kenichi Nakano, Hiroyuki Mori, Toshio Fukui1989 年 106 巻 4 号 p. 691-695
発行日: 1989年
公開日: 2011/01/25
ジャーナル フリーThe type L isozyme of potato tuber α-glucan phosphorylase [EC 2.4.1.1], a dimer of 104-kDa subunits, is compartmentalized in the amyloplast. We have cloned a nearly full-length cDNA encoding this isozyme from a cDNA library of immature potato tuber. The sequence was supplemented by a partial genomic clone. The transcription initiation site was identified by a primer extension experiment to be 43 bases upstream from the translation initiation ATG codon. The message encodes a polypeptide of 966 amino acid residues, of which 50 residues constitute an N-terminal extended peptide and 916 residues make up the mature protein. In the mature protein region, the nucleotide sequence is consistent with the chemically determined amino acid sequence (Nakano, K. & Fukui, T.(1986) J. Biol. Chem. 261, 8230-8236). The N-terminal extension bears characteristic features of the transit peptides of nuclear-encoded chloroplastic proteins, and is therefore regarded as a transit peptide for the amyloplast. This peptide is rich in basic amino acids (5 arginines, 3 lysines, and 5 histidines) and hydroxylic amino acids (7 serines and 5 threonines), but lacks acidic amino acids. It is therefore classified as one of the most basic transit peptides so far reported.抄録全体を表示PDF形式でダウンロード (2125K) -
A Cofilin-Like Protein Is Involved in the Regulation of Actin Assembly in Developing Skeletal MuscleHiroshi Abe, Sumiko Ohshima, Takashi Obinata1989 年 106 巻 4 号 p. 696-702
発行日: 1989年
公開日: 2011/01/25
ジャーナル フリーAn actin-binding protein of 20kDa (called 20K protein) was purified from the sarcoplasmic fraction of embryonic chicken skeletal muscle. The properties of this protein were very similar to cofilin, which was discovered in porcine brain (Nishida et al. (1984) Biochemistry, 23, 5307-5313): it bound to both G- and F-actin, inhibited actin polymerization in a pH-dependent manner, inhibited binding of tropomyosin to F-actin, and had almost the samemolecular size and pI as cofilin. A specific monoclonal antibody to 20K protein (MAB-22) was prepared to examine the expression and location of 20K protein during skeletal muscle development. When the whole protein lysates of embryonic and posthatched chicken skeletal muscles were examined by means of immunoblotting combinedwith SDS-PAGE, 20K protein was detected in skeletal muscle through the developmental stages. Location of 20K protein in the cells differed between the embryonic and adult tissues;immunofluorescence staining of the cryosections of embryonic muscle with MAB-22visualized irregular dot-like structures, but adult muscle sections were stained faintly and uniformly. 20K protein was present as a complex with actin in embryonic muscle, as judged by the ability to bind to a DNase I affinity column, while the same protein was free from actin in the cytoplasm of adult muscle. From these results, it is suggested that 20K protein regulates actin assembly transiently in developing skeletal muscle.抄録全体を表示PDF形式でダウンロード (4475K) -
Yoshihiko Sumi, Yataro Ichikawa, Yuichi Nakamura, Osamu Miura, Nobuo A ...1989 年 106 巻 4 号 p. 703-707
発行日: 1989年
公開日: 2011/01/25
ジャーナル フリーαs-Plasmin inhibitor (α2PI), one of the serine protease inhibitors in plasma, was expressed in baby hamster kidney (BHK) cell line. The expression vector was constructed with its genomic DNA and cDNA, and was transfected into BHK cells by the calcium phosphate method. The recombinant α2PI which was secreted from the cells was estimated by SDS-PAGE to have amolecular mass of 67kDa, which is indistinguishable from that of normal plasma α2PI. The leader peptide of 12 amino acids was retained at the amino terminus of the recombinant α2PI. This finding suggests that α2PI has pre-pro type processing and the propeptide of 12 amino acids is not removed in BHK cells. This pro-α2PI shows essentially the same inhibitory activity on plasmin and the same affinity for plasmin (ogen) as those of normal α2PI. However, the cross-linking ability to fibrin is reduced to less than one-third of that of normal α2PI. The cross-linking site is the glutamine residue located at the second position from the amino terminus of normal α2PI. The conformational change of this region caused by the addition of the propeptide may have affected the cross-linking capacity of the inhibitor.抄録全体を表示PDF形式でダウンロード (1690K) -
Koji Furuno, Toyoko Ishikawa, Kenji Akasaki, Shinji Yano, Yoshitaka Ta ...1989 年 106 巻 4 号 p. 708-716
発行日: 1989年
公開日: 2011/01/25
ジャーナル フリーWe have raised specific polyclonal immunoglobulin G (IgG) against a major lysosomal membrane sialoglycoprotein (LGP107) taken from rat liver and have prepared a conjugate of its Fab' fragment with horseradish peroxidase (HRP-anti LGP107 Fab') as a probe for the subcellular antigen. Electron immunocytochemistry in primary cultured rat hepatocytes showed that LGP107 resided primarily within lysosomes and was associated with luminal amorphous materials as well as limiting membranes. In addition, LGP107 was shown to be substantially distributed throughout the endocytic vacuolar system. The glycoprotein was found clustered in coated pits at the cell surface and localized along the surrounding membranes in endocytic vesicles. When cultured cells were exposed to HRP-anti LGP107 Fab', the antibody which was bound to its antigen within the coated pits was internalized via a system of endocytic vesicles and transported to lysosomes. During 20min of incubation at 37°C, the HRP tracer appeared at an early stage in small vesicles and movedprogressively to larger vesicles, including multivesicular bodies. After 1h, the tracer could be clearly seen in lysosomes heterogenous in shape and size. The existence of LGP107 in endocytic compartments and the uptake of and LGP107 antibody by hepatocytes were not blocked by prior treatment of the cells with cycloheximide and excess amounts of and LGP107 IgG. These data suggest that LGP107 circulates between the cell surface and lysosomes through the endocytic membrane traffic in hepatocytes.抄録全体を表示PDF形式でダウンロード (6946K) -
Koji Furuno, Shinji Yano, Kenji Akasaki, Yoshitaka Tanaka, Yasunori Ya ...1989 年 106 巻 4 号 p. 717-722
発行日: 1989年
公開日: 2011/01/25
ジャーナル フリーHRP-anti LGP107 Fab' and 125I-anti LGP107 IgG were used as probes to study the movement of LGP107 in the endocytic membrane transport system in primary cultured hepatocytes of rats. Following the addition of HRP-anti LGP107 Fab' to the culture medium, the transfer of the antibody conjugate from the cell surface to lysosomes was examined by cell fractionation on Percoll density gradients. The HRP tracer showed a bimodal subcellular distribution, in plasma membrane and lysosomal fractions. The amount of HRP found in the lysosomal fractions became larger as the period of cell incubation was increased. The rate of HRP accumulation in lysosomes was 0.13% of the administered load per hour per 106 cells. When cells were given 125I-anti LGP107 IgG, the antibody was not stored but was rapidly degraded in the lysosomes. The uptake of 125I-IgG by the cells, which was assessed by measuring the TCA-soluble radiolabeled degradation products released into the medium, increased proportionally to the administered concentration of the antibody and to the incubation time. The rate of uptake of the polyvalent 125I-IgG was comparable to that for the uptake of the monovalent HRP-Fab', and remained unchanged even after long exposure of the cells to a saturating concentraton of the polyvalent IgG. This uptake process continued for many hours in the cells exposed to the protein synthesis inhibitor, cycloheximide. These results suggest that there is a continuous circulation of LGP107 between the cell surface and lysosomes in hepatocytes.抄録全体を表示PDF形式でダウンロード (1241K) -
Akira Takeda, Shuichi Saheki, Takashi Shimazu, Nozomu Takeuchi1989 年 106 巻 4 号 p. 723-727
発行日: 1989年
公開日: 2011/01/25
ジャーナル フリーWe previously demonstrated that the 27-kDa major component protein in rat liver gap junctions was phosphorylated by protein kinase C in vitro (Takeda, A. et al.(1987) FEBS Lett. 210, 169-172). In this study, we examined this further and examined the phosphorylation of the 27-kDa gap junction protein in rat hepatocytes by metabolically labeling cells with [32P] orthophosphate and using a monoclonal antibody to immunoprecipitate the protein. The in vitro phosphorylation was inhibited by monoclonal antibodies recognizing the carboxyl-(C-) terminal domain of the 27-kDa protein. Protease digestion analysis revealed that phosphorylation occurred at the C-terminal domain. In rat hepatocytes, the phorbol esters, 12-O-tetradecanoylphorbol-13-acetate and phorbol-12, 13-dibutyrate, stimulated the 27-kDa protein phosphorylation, whereas 4α-phorbol-12, 13-didecanoate did not. 1-Oleoyl-2-acetyl-sn-glycerol also stimulated the 27-kDa protein phosphorylation. In addition, norepinephrine stimulated the phosphorylation and pretreatment of hepatocytes with staurosporine, a potent inhibitor of protein kinase C, inhibited this stimulatory effect of norepinephrine. Both in vitro and in hepatocytes, analysis of chemical cleavage of the 27-kDa phosphoprotein revealed that phosphorylation occurred mainly at a 10-kDa fragment which the antibodies recognized. These results indicate that protein kinase C phosphorylates the 27-kDa gap junction protein, not only in vitro but also in hepatocytes, at the C-terminal domain of the protein.抄録全体を表示PDF形式でダウンロード (2980K)
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