巻号一覧
83 巻, 6 号
選択された号の論文の33件中1～33を表示しています
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Miho TAKAHASHI, Tsuneko UCHIDA1978 年 83 巻 6 号 p. 1521-1532
発行日: 1978/06/25
公開日: 2008/11/18
ジャーナル フリーAn extracellular exonuclease has been found and purified about 10, 000-fold from the culture broth of an extreme thermophile, Thermus thermophilus HB8. The enzyme had an isoelectric point at pH 5.1 and seems to be of a multimolecular type, with molecular weights estimated to be ca. 530, 000 (Peak I) and around 330, 000 (Peak II) by gel filtration. The properties of the most highly purified enzyme fraction, Peak I were investigated. The enzyme requires divalent cations (Mg2+Sn2+>Ca2+, Mn2+) and is inactive in the presence of EDTA. The pH optimum is 9.4-9.5 in glycine-NaOH buffer and the optimum temperature is 85°C. The rate of hydrolysis increases in the order heat-denatured DNA>nativeDNA>RNA. The enzyme hydrolyzes deoxyoligonucleotides bearing 5'-monophosphate to liberate 5'-mononucleotides in an exonucleolytic manner. However, oligonucleotides lacking a 5'-phosphoryl group, irrespective of the presence or absence of phosphate at the 3'-termini, give both 5'-mononucleotides and dinucleoside monophosphates derived from the 5'-termini. It was also found that dinucleotides terminated with a 5'-phosphoryl group were cleaved to 5'-mononucleotides, but dinucleoside monophosphates were resistant to this enzyme. This exonuclease should be useful in the direct determination of sequence at the 5'-terminus and the penultimate position of oligonucleotides by the use of high-performance liquid chromatography.抄録全体を表示PDF形式でダウンロード (1246K) -
V. Optical Properties of Low Density LipoproteinsJun-ichi AZUMA, Naoki KASHIMURA, Tohru KOMANO1978 年 83 巻 6 号 p. 1533-1543
発行日: 1978/06/25
公開日: 2008/11/18
ジャーナル フリーThe circular dichroism (CD), optical rotatory dispersion (ORD), and fluorescence emission spectra of two subfractions of pig serum low density lipoproteins (LDL1 and LDL2) were compared. The contribution of the carbohydrate moiety to the CD and ORD spectra was estimated on the basis of data obtained from isolated glycopeptides and the constituent monosaccharides. The carbohydrate moiety had no effect on the conformation of the protein moieties of LDL1 and LDL2 (apoLDL1 and apoLDL2). However, the intensities of the observed extrema in the CD and ORD spectra of the glycopeptides were greater than those expected from the monosaccharide composition. This suggests the existence of secondary structure in the carbohydrate moiety. In contrast to the carbohydrate moiety, the contribution of the lipid moiety to the CD and ORD spectra could not be neglected. When the effect of the lipid moiety was subtracted from the CD and ORD spectra, the spectra due to apoLDL1 and apoLDL2 were quite similar. Delipidation in the presence of sodium dodecyl sulfate (SDS) induced an increase in the content of disordered structure and α-helix accompanied by a decrease in the β-structure. In the presence of SDS, marked quenching occurred in the fluorescence emission spectra with a blue shift of the maximum emission wavelength from 332 to 326 nm. ApoLDL1 and apoLDL2 showed quite similar SDS-induced conformational transitions. The secondary structures of apoLDL1 and apoLDL2 in the native lipoproteins were stable to changes of pH and temperature. However, this stability was lost in the presence of SDS. These results suggest the importance of the lipid moiety in maintaining the native secondary structures of LDL1 and LDL2. From the overall similarity of the optical properties of apoLDL1 and apoLDL2, we conclude that the secondary structures of apoLDL1 and apoLDL2 are identical.抄録全体を表示PDF形式でダウンロード (798K) -
Location and Role in Mitochondrial Protein DegradationYoshiko BANNO, Harold P. MORRIS, Nobuhiko KATUNUMA1978 年 83 巻 6 号 p. 1545-1554
発行日: 1978/06/25
公開日: 2008/11/18
ジャーナル フリー1. Hepatoma 8999 showed extremely high activity of serine protease, but similar activities of other lysosomal proteases to those of normal rat liver. 2. Serine protease from rat liver formed a single immunoprecipitation band against antiserum to purified protease from hepatoma 8999. 3. The serine proteases in rat liver and hepatoma 8999 were restricted to the inner membranes of the mitochondrial fraction. 4. Polyacrylamide gel electrophoresis with sodium dodecylsulfate showed that hepatoma 8999 mitochondria contained less of the slowest moving protein component than rat liver mitochondrial protein. This component was found to be the best substrate for mitochondrial serine protease in both liver and hepatoma 8999. 5. The role of serine protease in mitochondrial protein degradation is discussed on the basis of these results.抄録全体を表示PDF形式でダウンロード (2251K) -
II. Studies by Ultraviolet Difference Spectrophotometry and FluoritnetryTaro IZUMI, Hideo INOUE1978 年 83 巻 6 号 p. 1555-1566
発行日: 1978/06/25
公開日: 2008/11/18
ジャーナル フリーUltraviolet difference absorption spectra of lysozyme [EC 3. 2. 1. 17] at pH 2.1 produced by dioxane were measured as a function of dioxane concentration at 25°C. At 20-30% dioxane an abrupt decrease in molar difference absorption coefficients Δε was observed in plots of Δε292 and Δε285 vs. dioxane concentration; this reflects an unfolding process of lysozyme by dioxane. The magnitude of the decrease in Δε292 indicated the exposure of at least one buried tryptophyl residue to the solvent environment in this unfolding process. On the other hand, two linear portions were observed at 0-12% and 40-70% dioxane in the plots, corresponding to solvent effects of dioxane on native and denatured lysozyme, respectively. The number of exposed tryptophyl and tyrosyl residues in these dioxane concentration regions was estimated using the method described previously (Izumi & Inoue (1976) J. Biochem. 70, 1309-1321). The results showed that 3.9 tryptophyl and 1.5 tyrosyl residues were exposed at 0-12%. dioxane while 3.1 tryptophyl and 2.5 tyrosyl residues were exposed at 40-70% dioxane. Tryptophyl and tyrosyl exposures of denatured lysozyme at dioxane concentrations above 40% were also estimated on the basis of difference spectra obtained using a lysozyme solution containing 40% dioxane as a reference. In good agreement with the above results, this modified method gave an exposure of 3.4 tryptophyl and 2.3 tyrosyl residues for denatured lysozyme and some advantages of its use for denatured proteins are discussed. These results indicate that the tryptophyl behavior in denaturation is different from that of tyrosyl residues: the tyrosyl exposure is simply increased upon denaturation, while the tryptophyl exposure is increased in the unfolding process at 20-30% dioxane, and then decreased even below that in the native state above 40% dioxane, The decrease in tryptophyl exposure was attributed to intramolecular reorganization of lysozyme (with the disulfide bridges situated near the tryptophyl residues remaining intact) in the direction of a more helical conformation at higher dioxane concentrations.
Fluorescence spectra were also measured for lysozyme in aqueous dioxane mixtures at apparent pH 2.1-2.3 and 25°C. The results were consistent with the unfolding process observed by difference spectral measurements, but at high dioxane concentrations the fluorescence spectra of lysozyme behaved in a complicated manner. This appeared to reflect the state of the tryptophyl residues in relation to the conformational changes in lysozyme. Preliminary results on the sedimentation velocity behavior of lysozyme are also reported.抄録全体を表示PDF形式でダウンロード (1518K) -
Masashi KOSAKAI, Fumio YAMAUCHI, Zensaku YOSIZAWA1978 年 83 巻 6 号 p. 1567-1575
発行日: 1978/06/25
公開日: 2008/11/18
ジャーナル フリーPorcine heparin (α-heparin) and whale heparin (ω-heparin) were deaminated, and sulfated disaccharides, compounds b-2, e-2-1, e-2-2, and e-2-3 from the former and compounds b'-2 and e'-2-1 from the latter, were isolated from these products by high-voltage paper electrophoresis, gel filtration on Sephadex G-25 and column chromatography on Dowex 1×2 (Cl-form) in succession. Based on the results of chemical, GLC and p. m. r. spectral analyses, together with previous observations, the structures of these sulfated disaccharides are proposed to be as follows: compounds b-2 and b'-2, 4-O-(α-L-idopyranosyluronic acid 2-sulfate)-2, 5-anhydro D-mannose 6-sulfate; compounds e-2-1 and e'-2-1, 4-O-(α-L-idopyranosyluronic acid 2-sulfate)-2, 5-anhydro-n-mannose; compound e-2-2, 4-O-(β-n-glucopyranosyluronic acid)-2, 5-anhydroD-mannose 6-sulfate; compound e-2-3, 4-O-(α-L-idopyranosyluronic acid)-2, 5-anhydro-D-mannose 6-sulfate.
A high-voltage paper electrophoretogram of the deamination products of α-heparin was compared with a similar electrophoretogram of those of ω-heparin. Except for fraction a from the former, both deamination products were composed of seven corresponding fractions, though they differed strikingly in their proportions. The proportion of higher oligosaccharides to disaccharides was smaller in the case of α-heparin than in that of ω-heparin, reflecting the higher content of 2-acetamido-2-deoxy-D-glucose residues in the latter. The greatest difference was found in disaccharide fractions. The major fraction in α-heparin was disulfated disaccharide, compound b-2, whereas that in ω-heparin was monosulfated disaccharide, compound e'-2-1. The present results indicate a lower content of 6-O-sulfated 2-amino-2-deoxy-D-glucose in ω-heparin than in α-heparin. It is suggested therefore that a portion of 6-O-sulfates does not contribute significantly to the anticoagulant activity of heparin, since ω-heparin has a higher activity than a-heparin.抄録全体を表示PDF形式でダウンロード (1252K) -
Hirotomo YAMADA1978 年 83 巻 6 号 p. 1577-1581
発行日: 1978/06/25
公開日: 2008/11/18
ジャーナル フリーAminoacyl-tRNA synthetases of bakers' yeast (Saccharomyces cerevisiae) were adsorbed to a phosphocellulose (P-cellulose) column, and those specific for tyrosine [EC 6.1.1.1], threonine [EC 6.1.1.3], valine [EC 6.1.1.9], and isoleucine [EC 6.1.1.5] were eluted with several specific tRNAs. Elutions of these synthetases were affected by ATP and/or MgCl2. The effects of ATP and MgCl2 differ with synthetases. Elutions of tyrosyl- and valyl-tRNA synthetases with their cognate tRNAs were more specific in the presence of MgCl2. Isoleucyl-tRNA synthetase was eluted with its cognate tRNA in the presence of both ATP and MgCl2. On the other hand, threonyl-tRNA synthetase was eluted in the absence of ATP and MgCl2 with unfractionated tRNA but not with some non-cognate tRNAs. This suggests that elution of threonyl-tRNA synthetase is highly specific. The present data on the effects of ATP or MgCl2 or both on this affinity elution will be useful for simple and rapid purification of the synthetases.抄録全体を表示PDF形式でダウンロード (342K) -
Hirotomo YAMADA1978 年 83 巻 6 号 p. 1583-1589
発行日: 1978/06/25
公開日: 2008/11/18
ジャーナル フリーThreonyl-tRNA synthetase [EC 6.1.1.3] of Saccharomyces carlsbergensis was highly purified by a simple enzyme-substrate affinity method. After calcium-gel treatment of the crude enzyme extract, ammonium sulfate precipitation, and removal of polynucleotides with DEAE-cellulose, the enzyme was nonspecifically adsorbed onto phosphocellulose (P-cellulose), and then eluted specifically with a linear concentration gradient of threonine tRNA from torula yeast (Candida (Torulopsis) utilis).3 The enzyme was purified 303-fold from the calcium-gel supernatant. This purification method is very simple and time-saving. The purified enzyme gave a single band on SDS-gel electrophoresis, indicating a molecular weight of 82, 000, and exhibited a molecular weight of about 150, 000 by gel filtration. This suggests that the enzyme consists of two identical subunits. Some kinetic properties of the pure enzyme are described.抄録全体を表示PDF形式でダウンロード (909K) -
Reconstitution from Its SubunitsTakashi MATSUMOTO, Jiro TOBARI1978 年 83 巻 6 号 p. 1591-1597
発行日: 1978/06/25
公開日: 2008/11/18
ジャーナル フリーConditions for the restoration of catalytic activity from heavy and light subunits which had been isolated from methylamine dehydrogenase of Pseudomonas sp. J were investigated in vitro. Maximal restoration of the activity was obtained in 0.8M potassium phosphate buffer, pH 7.0-9.3, at 30°C with equimolar concentrations of the two subunits. Under the optimal conditions, the recovery of enzyme activity was 86%. and the time required for half-maximal recovery was about 3 min. Addition of bovine serum albumin or p-chloromercuribenzoate to the incubation mixture had no effect on the rate or extent of recovery of the enzyme activity. When the heavy subunit was added to the light subunit, the absorption spectrum of the light subunit changed to a form similar to that observed for the native enzyme. The concentration of methylamine required to change the spectrum of the light subunit was greatly decreased in the presence of the heavy subunit. Reconstituted enzyme was prepared from the isolated subunits and purified by gel chromatography. The reconstituted enzyme resembled the native enzyme in specific activity, molecular weight, substrate specificity, reaction mechanism, Michaelis constants for methylamine and phenazine methosulfate, susceptibility to inhibitor, and absorption, fluorescence and ESR spectra. However, it was less stable than the native enzyme to thermal and pH treatments. The CD spectrum was also slightly different from that of the native enzyme.抄録全体を表示PDF形式でダウンロード (973K) -
A Subunit EnzymeShigeki SHIRAI, Takashi MATSUMOTO, Jiro TOBARI1978 年 83 巻 6 号 p. 1599-1607
発行日: 1978/06/25
公開日: 2008/11/18
ジャーナル フリーA methylamine dehydrogenase was purified to homogeneity from Pseudomonas AMI and obtained in crystalline form. It was found to be a subunit enzyme composed of two kinds of subunit, light and heavy. These two subunits were isolated by Sephadex G-100 gel chromatography after incubation of the enzyme with guanidine hydrochloride. Molecular weights of 13, 000 daltons for the light subunit and 40, 000 daltons for the heavy subunit were estimated by SDS-polyacrylamide gel electrophoresis, and the molecular weight of the native enzyme was found to be 105, 000 daltons by Sephadex G-200 gel chromatography. The enzyme and its subunits were also analyzed for amino acid composition, isoelectric point, and absorption, fluorescence, and CD spectra, as well as for the effects of pH, thermal treatment, and guanidine hydrochloride treatment. Both the subunits were absolutely required for enzymatic activity, either subunit alone being inactive. It could be deduced from the absorption and fluorescence spectra of the subunits that the prosthetic group of the enzyme was bound solely to the light subunit. These results suggest that the enzyme is a subunit enzyme similar to that of Pseudomonas sp. J, of the α2β2 type.抄録全体を表示PDF形式でダウンロード (1776K) -
Agglutination of Free and Membrane-Bound Ribosomes from Wheat Germ by Concanavalin AKazuo YOSHIDA1978 年 83 巻 6 号 p. 1609-1614
発行日: 1978/06/25
公開日: 2008/11/18
ジャーナル フリーFree and membrane-bound ribosomes prepared from wheat germ and extensively washed with 1% Triton X100-sodium deoxycholate, formed large aggregates in the presence of concanavalin A. The ribosomal aggregates were detected by sucrose density gradient centrifugation and immunoprecipitation. SDS-polyacrylamide gel electrophoresis of ribosomal proteins indicated that four ribosomal glycoproteins (MW about 14, 000 to 24, 000) were present in both free and membrane-bound ribosomes. However, three glycoproteins (MW about 25, 000 to 15, 000) appear to be underglycosylated or lacking in free ribosomes.抄録全体を表示PDF形式でダウンロード (1566K) -
Mutant Cyclic AMP Receptor Protein with an Increased Affinity for Cyclic AMPYutaka TAKEBE, Masabumi SHIBUYA, Yoshito KAZIRO1978 年 83 巻 6 号 p. 1615-1623
発行日: 1978/06/25
公開日: 2008/11/18
ジャーナル フリーA strain bearing an extragenic suppressor of cya mutation was isolated as a second-site revertant of an adenylate cyclase deficient strain. The mutant was unable to synthesize cAMP but showed normal fermentation profiles and growth properties on a variety of carbon sources. The site of reversion was mapped in, or near, the structural gene for the cAMP receptor protein. Structural alteration of the protein was directly demonstrated by the following biochemical observations: (i) A 10-fold decrease in the dissociation constant for cAMP, (ii) an acidic shift in the isoelectric point, and (iii) the altered binding properties to λh80dlac ps DNA.抄録全体を表示PDF形式でダウンロード (1249K) -
Kazutosi NISIZAWA, Takahisa KANDA, Shogo SHIKATA, Kazumasa WAKABAYASHI1978 年 83 巻 6 号 p. 1625-1630
発行日: 1978/06/25
公開日: 2008/11/18
ジャーナル フリーMutarotation of products from p-nitrophenyl β-n-cellobioside and cellopentaitol by two different types of exo-cellulases from Trichoderma viride was investigated. It was found that an exo-cellulase of glucosidase type produced from the former substrate D-glucose which was mutarotated in a downward direction, while another exo-cellulase of Avicelase type produced from the latter substrate cellobiose which was mutarotated in an upward direction.抄録全体を表示PDF形式でダウンロード (1058K) -
I. Isolation and Characterization of CytochromesKen-ichiro SHIMAZAKI, Ken-ichiro TAKAMIYA, Mitsuo NISHIMURA1978 年 83 巻 6 号 p. 1631-1638
発行日: 1978/06/25
公開日: 2008/11/18
ジャーナル フリーTwo cytochromes of the C type, c-550 and c-553, were isolated from the marine diatom, Phaeodactyhum tricornutum, and purified by ammonium sulfate fractionation and DEAEcellulose column chromatography.
The cytochrome c-550 had absorption maxima at 550, 522, and 417nm in the reduced form and at 524, 407, 351, and 277nm in the oxidized form. It was an autoxidizable acidic protein with an isoelectric point of 5.1 and had a low redox potential of about -0.20V at pH 7.0. The molecular weight of this cytochrome was close to 17, 000. This cytochrome combined with CO and CN-. The CO complex was dissociated reversibly by light.
The cytochrome c-553 had absorption maxima at 553, 522.5, and 417nm in the reduced form and at 528, 410, and 356nm in the oxidized form. The protein had an acidic isoelectric point of 3.7 and had a high mid-point redox potential of +0.36V at pH 7.0. Its molecular weight was approximately 10, 500. The cytochrome may be considered to be a photosynthetic cytochrome of the f type.
Cytochromes of the B type were also found in Phaeodactylum tricornutum; one in soluble form, and the other in bound form. The soluble form had absorption maxima at 560, 529, and 427nm in the reduced state and at 413nm in the oxidized state.抄録全体を表示PDF形式でダウンロード (482K) -
II. Identification and Determination of Quinones, Cytochromes, and FlavinsKen-ichiro SHIMAZAKI, Ken-ichiro TAKAMIYA, Mitsuo NISHIMURA1978 年 83 巻 6 号 p. 1639-1642
発行日: 1978/06/25
公開日: 2008/11/18
ジャーナル フリーQuinones constituting the electron transfer systems in a marine unicellular diatom, Phaeodactylum tricornutum, were isolated and identified chromatographically. The alga contained five quinones, i. e., plastoquinone A, plastoquinone C, plastoquinone D, α-tocopherylquinone, and ubiquinone-9. Other types of quinones, such as vitamin K1, were not detected. The contents of plastoquinone A, plastoquinone C, plastoquinone D, ubiquinone-9, and α-tocopherylquinone were 25.5, 4.95, 1.99, 4.78, and 0.28 mmol per mol of chlorophyll, respectively.
The contents of the soluble C-type cytochromes, cytochrome c-550 and cytochrome c-553, were 2.15 and 4.34 mmol (heme basis) per mol of chlorophyll, respectively. The amount of B-type cytochrome in the bound form was estimated to be 3.24 mmol (heme basis) per mol of chlorophyll. The acid-soluble flavins, FAD and FMN, were present in amounts of 0.68 and 0.41 mmol per mol of chlorophyll, respectively.抄録全体を表示PDF形式でダウンロード (286K) -
Mitsuo IKEBE, Toshimitsu AIBA, Hirofumi ONISHI, Shizuo WATANABE1978 年 83 巻 6 号 p. 1643-1655
発行日: 1978/06/25
公開日: 2008/11/18
ジャーナル フリーActin, myosin, and “native” tropomyosin (NTM) were separately isolated from chicken gizzard muscle and rabbit skeletal muscle. With various combinations of the isolated contractile proteins, Mg-ATPase activity and superprecipitation activity were measured. It was thus found that gizzard myosin and gizzard NTM behaved differently from skeletal myosin and skeletal NTM, whereas gizzard actin functioned in the same way as skeletal actin. It was also found that gizzard myosin preparations were often Ca-sensitive, that is, that the two activities of gizzard myosin plus actin without NTM were activated by low concentrations of Ca2+. The Mg-ATPase activity of a Ca-insensitive preparation of gizzard myosin was not activated by actin even in the presence of Ca2+.
When Ca-sensitive gizzard myosin was incubated with ATP (and Mg2+) in the presence of Ca2+, a light-chain component of gizzard myosin was phosphorylated. The light-chain phosphorylation also occurred when Ca-insensitive myosin was incubated with gizzard NTM and ATP (plus Mg2+) in the presence of Ca2+. In either case, the light-chain phosphorylation required Ca2+. Phosphorylated gizzard myosin in combination with actin was able to exhibit superprecipitation, and Mg-ATPase of the phosphorylated gizzard myosin was activated by actin; the actin activation and superprecipitation were found to occur even in the absence of Ca2+ and NTM or tropomyosin.
The phosphorylated light-chain component was found to be dephosphorylated by a partially purified preparation of gizzard myosin light-chain phosphatase. Gizzard myosin thus dephosphorylated behaved exactly like untreated Ca-insensitive gizzard myosin; in combination with actin, it did not superprecipitate either in the presence of Ca2+ or in its absence, but did superprecipitate in the presence of NTM and Ca2+.
Ca-activated hydrolysis of ATP catalyzed by gizzard myosin B proceeded at a reduced rate after removal of Ca2+ (by adding EGTA), whereas that catalyzed by a combination of actin, gizzard myosin, and gizzard NTM proceeded at the same rate even after removal of抄録全体を表示PDF形式でダウンロード (1958K) -
Toshiharu HASE, Sadao WAKABAYASHI, Hiroshi MATSUBARA, Lorenz KERSCHER, ...1978 年 83 巻 6 号 p. 1657-1670
発行日: 1978/06/25
公開日: 2008/11/18
ジャーナル フリー1. The complete amino acid sequence of the 2Fe-2S ferredoxin from Halobacterium halobium was determined to be:
Pro-Thr-Val-Glu-Tyr-Leu-Asn-Tyr-Glu-Thr-Leu-Asp-Asp-Gln-Gly-Trp-Asp-Met-Asp-Asp-Asp-Asp-Leu-Phe-Glu-Lys-Ala-Ala-Asp-Ala-Gly-Leu-Asp-Gly-Glu-Asp-Tyr-Gly-Thr-Met-Glu-Val-Ala-Glu-Gly-Glu-Tyr-Iie-Leu-Glu-Ala-Ala Glu-Ala-Gln-Gly-Tyr-Asp-Trp-Pro-Phe-Ser-Cys-Arg-Ala-Gly-Ala-Cys-Ala-Asn- Cys-Ala- Ser-Ile- Val-Lys- Glu- Gly- Glu-Ile-Asp-Met-Asp-Met-Gin -Gin -Tie- Leu-Ser-Asp -Glu-Glu-Val-Glu-Glu-Lys-Asp -Val-Arg-Leu-Thr-Cys-Ile-Gly-Ser-Pro-Ala-Ala-Asp-Glu- Val-Lys-Ile- Val-Tyr-Asn-Ala-Lys-His-Leu-
Ac
Asp-Tyr-Leu-Gln-Asn-Arg-Val-Ile
2. The apoferredoxin chain consists of 128 amino acid residues and has a molecular weight of 14, 330.
3. There are only four cysteines in this ferredoxin molecule; they should be involved in the binding of the two iron atoms at the active center. The relative positions of these cysteines are similar to those of the cysteines in chloroplast ferredoxins.
4. There is a high degree of homology between H. halobium ferredoxin and chloroplast ferredoxins, though the latter molecules contain only about 98 amino acid residues.
5. H. halobium ferredoxin contains a single residue of Nε-acetyllysine.抄録全体を表示PDF形式でダウンロード (796K) -
Toshiharu HASE, Hiroshi MATSUBARA, Hajime YOSHIZUMI1978 年 83 巻 6 号 p. 1671-1678
発行日: 1978/06/25
公開日: 2008/11/18
ジャーナル フリーPurothionin isolated from commercial wheat flour contained several components and two of them (A-I and A-II) were isolated in pure form by CM-52 column chromatography. Each component contained 45 amino acid residues with 4 disulfide bonds. Purothionin A-11 was digested with trypsin and thermolysin to isolate cystine peptides. These were separated and purified by chromatography on an SP-Sephadex column, and paper electrophoresis and chromatography. A peptide containing a -Cys-Cys- sequence was hydrolyzed with 10 N sulfuric acid. Amino acid compositions and partial sequence studies of the cystine peptides and their performic acid-oxidized peptides revealed the positions of all 4 disulfide bonds in purothionin A-II. They were formed between residues 3 and 39, 4 and 31, 12 and 29, and 16 and 25. The results of a partial study of purothionin A-I are also presented.抄録全体を表示PDF形式でダウンロード (515K) -
Jun MIYAKE, Sonoe OCHIAI-YANAGI, Takafumi KASUMI, Toshio TAKAGI1978 年 83 巻 6 号 p. 1679-1686
発行日: 1978/06/25
公開日: 2008/11/18
ジャーナル フリーA membrane protein insoluble in water was isolated by gel chromatography in the presence of 0.1% sodium dodecyl sulfate (SDS) from chromatophores of a photosynthetic bacterium, Rhodospirillum rubrum. This is one of the major membrane proteins of the chromatophore. The protein was found to bind about four grams of SDS per gram, a value which is more than twice the amount generally observed with protein polypeptides derived from watersoluble globular proteins.
The electrophoretic behavior of the complex between the membrane protein and SDS is abnormal due to this high capacity for binding SDS. Estimation of the molecular weight of this protein by SDS-polyacrylamide gel electrophoresis was thus impossible. Such an anomaly in SDS binding is unlikely to be restricted to the particular membrane protein described in this paper. The possibility of such a deviation from standard behavior in the interaction with SDS should be taken into consideration in studies of other membrane proteins, since SDS is often used both in analytical and preparative procedures.抄録全体を表示PDF形式でダウンロード (1278K) -
I. Detection of the Phase Transition and Its Dependence on Growth TemperatureNobuyuki WAKAYAMA, Tairo OSHIMA1978 年 83 巻 6 号 p. 1687-1692
発行日: 1978/06/25
公開日: 2008/11/18
ジャーナル フリーPhase transition was detected by a fluorescence polarization technique in the membrane of Thermus thermophilus HB8: it was found to be a function of cell growth temperature when the cell growth temperature was varied between 50-80°C.
A systematic relation between the phase transition temperature and the growth temperature was observed. Differential scanning calorimetry was also applied. The phase transition was found to take place between 34 and 55°C for cells grown at 50°C and between 52 and 80°C for cells grown at 80°C.抄録全体を表示PDF形式でダウンロード (367K) -
II. Membrane Functions Underlying Leucine Transport and Their Relation with Thermotropic Phase TransitionsNobuyuki WAKAYAMA1978 年 83 巻 6 号 p. 1693-1698
発行日: 1978/06/25
公開日: 2008/11/18
ジャーナル フリーVarious membrane properties of Thermus thermophilus HB8 were studied in order to elucidate the mechanism and implications of cell adaptation to a high temperature environment.
1. The profile of temperature dependence of the amino acid uptake rates was similar for the membrane obtained from cells grown at both 61°C and 77°C.
2. The remaining uptake activity after heat treatment paralleled the membrane potential generating ability, as measured by auramine O fluorescence response to the respiration substrate, and was not dependent on the growth temperature.
3. The temperature dependence of the efflux of the accumulated amino acid reflects the physical phase of the membrane and is dependent on the growth temperature.抄録全体を表示PDF形式でダウンロード (359K) -
Junko SOKAWA, Yoshihiro SOKAWA1978 年 83 巻 6 号 p. 1699-1705
発行日: 1978/06/25
公開日: 2008/11/18
ジャーナル フリーIn 10B601 (rel+) strain possessing a temperature-sensitive valyl-tRNA synthetase, chloramphenicol prevented the formation of guanosine-3'-diphosphate-'-diphosphate (ppGpp) as well as the stringent control of stable RNA synthesis, under the conditions where the incorporation of valine into protein was still detectable, i.e. at the lower restrictive temperatures. On the other hand, the effect of chloramphenicol was not observed at higher restrictive temperatures above 42°C where the incorporation of valine was completely absent.
Pretreatment of 10B601 cells with chloramphenicol before transfer to a high restrictive temperature (43.5°C) did retard the onset of accumulation of ppGpp after the shift-up. Duration of the lag period was dependent on the concentration of chloramphenicol added. In parallel with the inability of the cells to accumulate ppGpp, stable RNA synthesis was permitted to continue at that high temperature.
These results suggest that chloramphenicol traps aminoacyl-tRNA at the A-sites of ribosomes by damming-up the small flow of aminoacyl-tRNA under the restrictive supply of amino acids. Uncharged tRNA which has been located at the A-site is replaced by the charged one, thus resulting in the suppression of ppGpp formation and in the restoration of stable RNA synthesis.抄録全体を表示PDF形式でダウンロード (522K) -
The Wavelength Dependence of Observed Oxygen Equilibrium Parameter ValuesKazuhiko IMAIZUMI, Kiyohiro IMAI, Itiro TYUMA1978 年 83 巻 6 号 p. 1707-1713
発行日: 1978/06/25
公開日: 2008/11/18
ジャーナル フリーSpectral changes of oxyhemoglobin induced by such anions as 2, 3-diphosphoglycerate, inositol hexaphosphate, and Cl- may affect the validity of the spectrophotometric determination of oxygen saturation of hemoglobin. Therefore, the anion-induced difference spectra were extensively measured under a variety of conditions and accurate oxygen equilibrium curves were determined under representative conditions with detection at different wavelengths selected from peaks, troughs, and zero difference points of the difference spectra in the visible and Soret regions. Oxygen equilibrium parameters including the four Adair constants (i.e., equilibrium constants for four steps of oxygenation) estimated from the equilibrium curves did not show any dependence on wavelength within the limits of experimental error. These results indicate that anion-induced spectral changes do not invalidate the spectrophotometric determination of oxygen saturation and confirm the validity of the previous conclusions drawn in our series of studies on the effects of anions, pH and temperature on oxygen equilibrium parameters.抄録全体を表示PDF形式でダウンロード (467K) -
Sigehiro MORITA, Tatsuo MIYAZAKI1978 年 83 巻 6 号 p. 1715-1720
発行日: 1978/06/25
公開日: 2008/11/18
ジャーナル フリーThe dichroism was measured in films of air-dried and, consequently, flattened chromatophores of Chromatium vinosum, Rhodopseudomonas sphaeroides and Rhodospirillum rubrum. The values (ΔA/A) of dichroism in C. vinosum were found to be -1.05 at 590nm and 0.75 in the near infrared region. The values of dichroism in R. sphaeroides were -0.70 at 590nm and 0.80 at 870nm. The values of dichroism in R. rubrum were -1.45 at 590nm and 0.97 at 870nm.抄録全体を表示PDF形式でダウンロード (439K) -
III. Purification and Properties of Bovine Glucose 1, 6-Bisphosphate SynthaseMasatsugu UEDA, Masaaki HIROSE, Ryuzo SASAKI, Hideo CHIBA1978 年 83 巻 6 号 p. 1721-1730
発行日: 1978/06/25
公開日: 2008/11/18
ジャーナル フリーGlucose 1, 6-bisphosphate synthase, which catalyzes the synthesis of glucose 1, 6-bisphosphate from 1, 3-bisphosphoglycerate and glucose monophosphates, was partially purified from bovine liver and bovine red cells. The liver synthase was separated into two peaks by Blue dextran Sepharose 4B column chromatography (peak I and peak II). The peak I and the peak II enzyme both appeared to utilize glucose 1-phosphate, glucose 6-phosphate and mannose 1-phosphate as phosphate acceptors. With respect to phosphate donor, these enzyme were specific for 1, 3-bisphosphoglycerate.
The Km values for 1, 3-bisphosphoglycerate were very similar for the peak I and peak II enzymes (peak I, 0.23μM; peak II, 0.28μM;). By contrast, the Km values for glucose monophosphates were significantly different for these two peaks (peak I; KmGlc-1-P 9μM, KmGtc-s-P 15μM: peak II; KmGlc-1-P 59μM, KmGlc-s-P 61μM). The Km values of the red cell synthase were greater than those of the liver synthases (KmGlc-l-P, 160μM KmGtycerate-1, 3-P2, 0.45μM), suggesting that the red cell synthase is a different enzyme species from the liver synthase.
Bovine liver glucose 1, 6-bisphosphate synthases were strongly inhibited by fructose 1, 6-bisphosphate and 2, 3-bisphosphoglycerate competitively with respect to 1, 3-bisphosphoglycerate.
Bovine liver phosphoglucomutase also catalyzes the synthesis of glucose 1, 6-bisphosphate, and this activity is affected by the bisphosphate compounds (Hirose, M., Ueda, M., & Chiba, H. (1976) Agric. Biol. Chem. 40, 2433-2439). Glucose 1, 6-bisphosphate synthesizing activities of the synthases and phosphoglucomutase were compared in a liver homogenate based on the kinetic constants of these enzymes. It is suggested that the synthases were predominant in the presence of the normal level of glucose 1, 6-bisphosphate, whereas phosphoglucomutase acted as a glucose 1, 6-bisphosphate synthesizing enzyme only in the presence of an abnormally low level of glucose 1, 6-bisphosphate.抄録全体を表示PDF形式でダウンロード (721K) -
Jun NAKAMURA, Kazuhiko KONISHI1978 年 83 巻 6 号 p. 1731-1735
発行日: 1978/06/25
公開日: 2008/11/18
ジャーナル フリーThe effects of K+ and Na+ on the Ca2+, Mg2+-ATPase of sarcoplasmic reticulum fragments (SRF) were investigated at 1mM ATP. There was an alteration of the sensitivity of the ATPase to the monovalent cations during storage of the SRF preparation.
The Ca2+, Mg2+-ATPase of freshly prepared SRF was slightly activated by 5-10mM K+ and Na+. Mg2+-ATPase was inhibited by both the monovalent cations to the same extent, and this response to the ions was independent of the freshness of the preparations. After storage of SRF, however, the Ca2+, Mg2+-ATPase was markedly activated by higher concentrations of K+ and Na+ (0.2-0.3M).
K+ and Na+ reduced the Ca uptake at the steady state in freshly prepared SRF, but did not affect pre-steady state uptake. In the presence of oxalate, the rate of Ca accumulation both in fresh and stored preparations was activated by 0.1-0.2M K+ and Na+. The Ca2+, Mg2+-ATPase with oxalate, so-called “extra ATPase, ” showed the same response to the ions as did the activity without oxalate during storage.抄録全体を表示PDF形式でダウンロード (376K) -
Isolation, Characterization, and Chemical ModificationOkitoshi ABE, Junko OHATA, Yoko UTSUMI, Kenji KUROMIZU1978 年 83 巻 6 号 p. 1737-1748
発行日: 1978/06/25
公開日: 2008/11/18
ジャーナル フリーA proteinase inhibitor which inhibits trypsin [EC 3. 4. 21. 4] and α-chymotrypsin [EC 3. 4. 21. 1] was isolated from the seeds of Vicia angustifolia L. var. segetalis Koch and purified by gel filtration on Sephadex G-100 followed by ion-exchange chromatography on DEAE-Sephadex A-25. The inhibitor has a molecular weight of about 13, 000 when determined by molecularsieve chromatography on Sephadex G-100 and 12, 600 from the amino acid composition, which revealed high contents of half-cystine, aspartic acid, serine, and glutamic acid residues. The absence of free sulfhydryl groups, tryptophan, and carbohydrate was also observed. The amino- and carboxyl-terminal residues of the inhibitor were determined to be glycine and asparagine, respectively, suggesting that this inhibitor is composed of a single polypeptide chain. The inhibitor has an isoelectric point at pH 7.6 and is considerably stable in acidic and neutral pH regions, however, it loses inhibitory activities at alkaline pH.
Stoichiometry of the reactions between the inhibitor and the enzymes showed that this inhibitor reacted with trypsin in a 1:1 and with chymotrypsin in a 1:2 molar ratio.
Maleylation of the inhibitor revealed that even when all the 9 amino groups, i.e. one α-amino and 8 ε-amino groups, were modified, the inhibitor retained its full antitryptic activity. On the other hand, on 1, 2-cyclohexanedione treatment, the inhibitor lost almost all the antitryptic activity and also 50% of the antichymotryptic activity. Modifications of all the three tyrosyl residues in the inhibitor decreased the antichymotryptic activity by 50% but had no effect on the antitryptic activity. The results of these chemical modifications seem to support that this inhibitor has one reactive site for trypsin and two for α-chymotrypsin.抄録全体を表示PDF形式でダウンロード (821K) -
Okitoshi ABE, Yukiko SHIMOKAWA, Tomiko ARAKI, Kenji KUROMIZU1978 年 83 巻 6 号 p. 1749-1756
発行日: 1978/06/25
公開日: 2008/11/18
ジャーナル フリーTrypsin [EC 3. 4. 21. 4] modified (reactive site cleaved) Vicia angustifolia proteinase inhibitor was prepared at pH 3 with a catalytic amount of trypsin and purified using columns of Sephadex G-50 and DEAE-Sephadex A-25. The modified inhibitor, which still retained antitryptic activity, lost its activity upon treatment with carboxypeptidase B or citraconic anhydride. End-group analyses revealed that the carboxyl-terminal Arg and the amino-terminal Ser residues were newly exposed end-groups in the modified inhibitor. It takes a much longer incubation time (about 1h) to exhibit the maximal inhibitory activity against trypsin. Reduction and carboxymethylation of the modified inhibitor produced two fragments on Sephadex G-50 chromatography. The smaller fragment consisted of about 32 amino acid residues and possessed a new carboxyl-terminal Arg residue. The larger fragment consisted of about 80 residues and possessed a Ser residue at its amino-terminus. These results indicate that the small fragment was derived from the amino-terminal portion of the modified inhibitor and the large fragment from the carboxyl-terminal. It is also concluded that an Arg-Ser bond is the reactive site as well as the inhibitory site of the V. angustifolia inhibitor against trypsin. The sequence around the antitryptic site exhibits high degrees of homology with other double-headed inhibitors of legume origin, such as the Bowman-Birk inhibitor, lima bean inhibitor, and the major inhibitor in chick-peas.抄録全体を表示PDF形式でダウンロード (1019K) -
Tadashi ISHIMODA-TAKAGI1978 年 83 巻 6 号 p. 1757-1762
発行日: 1978/06/25
公開日: 2008/11/18
ジャーナル フリーThe antiserum against lantern muscle tropomyosin of the sea urchin was prepared, and the presence of tropomyosin in the sea urchin egg was shown by immunodiffusion test between the antiserum and the egg tropomyosin fraction which was prepared according to the purification method for muscle tropomyosin. The sea urchin egg tropomyosin was isolated from the immuno-precipitate formed between the antiserum and the egg tropomyosin fraction. The subunit molecular weight of the egg tropomyosin was calculated to be 29, 000.抄録全体を表示PDF形式でダウンロード (1422K) -
Ken-ichi KODAIRA, Akira TAKETO1978 年 83 巻 6 号 p. 1763-1767
発行日: 1978/06/25
公開日: 2008/11/18
ジャーナル フリーHost dna functions involved in the replication of microvirid phage øC DNA were investigated in vivo. Although growth of this phage was markedly inhibited even at 35-37°C even in dna+ host, conversion of the infecting single-stranded DNA into the double-stranded parental replicative form (stage I synthesis) occurred normally at 43°C in dna+, dnaA, dnaB, dnaC (D), and dnaE cells. In dnaG mutant, the stage I synthesis was severely inhibited at 43°C but not at 30°C. The stage I replication of øC DNA was clearly thermosensitive in dnaZ cells incubated in nutrient broth. In Tris-casamino acids-glucose medium, however, the dnaZ mutant sufficiently supported synthesis of the parental replicative form. At 43°C, synthesis of the progeny replicative form DNA (stage II replication) was significantly inhibited even in dna+ cells and was nearly completely blocked in dnaB or dnaC (D) mutant. At 37°C, the stage II replication proceeded normally in dna+ bacteria.抄録全体を表示PDF形式でダウンロード (355K) -
I. High-Affinity Binding SitesGlen Emery HOFMANN, William Stanley GLAUNSINGER1978 年 83 巻 6 号 p. 1769-1778
発行日: 1978/06/25
公開日: 2008/11/18
ジャーナル フリーThe high-affinity (n1, ) Mn (II) binding sites in glutamine synthetase [EC 6.3.1.2] have been examined as a function of Mn (II) concentration and the state of adenylylation by electron paramagnetic resonance (EPR) spectroscopy. Mn (II) EPR spectra are characteristic of paramagnetically dilute samples, and studies of the Mn(II) concentration dependence has indicated that these cations remain magnetically isolated (interaction distance_??_15-17Å) both at different states of adenylylation and in the presence of the inhibitor L-methionine (SR)-sulfoximine (MSOX). An investigation of metal-enzyme complexes at various states of adenylylation has revealed that the n1, Mn (II) sites are not perturbed by the addition of 5'-adenylyl groups, suggesting that the n1, Mn (H) sites are well removed from these groups. The small axial distortion (_??_15G) in these complexes is indicative of a nearly isotropic environment for Mn (II), and the invariance of linewidth with adenylylation and incorporation of L-glutamate suggests that the dominant relaxation mechanism for Mn (II) may not involve collisions with solvent molecules. In the presence of MSOX, the linewidth narrows drastically and forbidden transitions are observed. The n1, Mn (II) site experiences an axial distortion which decreases linearly with increasing adenylylation. In addition, the Mn (II) hyperfine coupling constant, and hence bond ionicity, increases linearly with adenylylation. These results are interpreted in terms of possible longitudinal and transverse distortion mechanisms and their implications for catalysis are discussed. The measured conformational and bonding changes at the n1, Mn (II) sites may reflect corresponding changes near the γ-carboxyl group of L-glutamate, whose activation is believed to be required for the biosynthetic reaction.抄録全体を表示PDF形式でダウンロード (778K) -
II. Intermediate-Affinity Binding SitesGlen Emery HOFMANN, William Stanley GLAUNSINGER1978 年 83 巻 6 号 p. 1779-1782
発行日: 1978/06/25
公開日: 2008/11/18
ジャーナル フリーThe nature of the intermediate-affinity (n2) Mn (II) binding sites in glutamine synthetase [EC 6.3.1.2] has been studied as a function of adenylylation in a variety of enzyme-metal complexes by EPR. In the absence of nucleotide, the n2 Mn (II) environment is nearly isotropic, the Mn (II) bonds are highly ionic, and the interaction distance R≥12-14A. Nucleotide binding at the n2 Mn (II) site renders the n2 Mn (II) signal unobservable and causes a reduction in signal amplitude (_??_30%) and line broadening (_??_6G) at the high-affinity (n1) Mn (II) site. This behavior indicates that nucleotide binding induces a conformational change in the enzyme which brings the previously distant nl and n2 sites into closer proximity (RG≤8-11A), possibly for the purpose of activating the nucleotide for direct phosphoryl transfer to L-glutamate. In line with this suggestion, the broad, unresolved resonances in complexes containing both L-methionine SR-sulfoximine (MSOX) and nucleotide may result from the phosphorylation of MSOX. The n2 Mn (II) site is not affected by adenylylation in all the enzyme-metal complexes studied, which suggests that the regulatory effects of adenylylation may only act at the n1 Mn (II) sites.抄録全体を表示PDF形式でダウンロード (286K) -
Kazuhiro SOGAWA, Kenji TAKAHASHI1978 年 83 巻 6 号 p. 1783-1787
発行日: 1978/06/25
公開日: 2008/11/18
ジャーナル フリーConditions have been investigated for the use of fluorescamine-labeled casein as a substrate for fluorometric assay of proteinases. Fluorescamine-labeled casein can be prepared simply by mixing solutions of casein and fluorescamine at pH 8.0 and used without removal of the excess reagent or its hydrolysis product. The fluorescence of the labeled casein and its enzymatic digest is moderately stable in the range of pH 7.0 to 10.0. Activities can be determined by measuring the fluorescence of the hydrolysis products soluble in 0.1M trichloroacetic acid solution at pH 4.0 after adjusting the pH of the acid-soluble fraction to 7.7. This method is suited for assay of proteinases active at neutral to slightly alkaline pH values, and is capable of quantitating about 0.05μg of trypsin or 0.5μg of α-chymotrypsin or papain. The assay can be done in the presence of large amounts of contaminating amino acid, protein and/or exopeptidases which may interfere with the ordinary assay of proteinases.抄録全体を表示PDF形式でダウンロード (371K) -
VI. Surface Charge of Very Low Density LipoproteinJun-ichi AZUMA, Tohru KOMANO1978 年 83 巻 6 号 p. 1789-1791
発行日: 1978/06/25
公開日: 2008/11/18
ジャーナル フリーThe surface electric charge of pig serum very low density lipoprotein (VLDL) is described. By isoelectric focusing VLDL was separated into at least 3 fractions having different isoelectric points and polypeptide distributions. The ultracentrifugal and electron microscopic results indicate that the VLDL was not drastically denatured by Ampholine.抄録全体を表示PDF形式でダウンロード (700K)
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