巻号一覧
92 巻, 1 号
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Yoshikazu ICHIHARA, Kazuhiro SOGAWA, Kenji TAKAHASHI1982 年 92 巻 1 号 p. 1-5
発行日: 1982年
公開日: 2008/11/18
ジャーナル フリーA simple and sensitive method for proteinase assay was developed, which uses fluorescamine-labeled casein-Sepharose 4B as a substrate. Casein-Sepharose 4B was prepared most effectively by coupling casein to cyanogen bromide-activated Sepharose 4B at pH 10.0. Fluorescamine-labeled casein-Sepharose 4B was then prepared by mixing fluorescamine and casein-Sepharose 4B suspension at pH 8.0 and used for the assay as a substrate after removal of the excess reagent and/or its hydrolysis products. The assay can be done by simply measuring the fluorescence (excitation at 390 nm and emission at 475 nm) of the filtrate of the assay mixture after incubation of the substrate with enzyme solution.
This method is suited for the assay of proteinases active at neutral to slightly alkaline pH values, and the activity of 3 ng of trypsin or 10 ng of α-chymotrypsin can be determined with reasonable accuracy. This method is therefore almost as sensitive as those using radioisotope-labeled proteins as substrates.抄録全体を表示PDF形式でダウンロード (377K) -
Akihiko KAWAGUCHI, Hiroshi TOMODA, Shigeo NOZOE, Satoshi OMURA, Shigen ...1982 年 92 巻 1 号 p. 7-12
発行日: 1982年
公開日: 2008/11/18
ジャーナル フリーCerulenin, an antibiotic with the structure of (2R) (3S)-2, 3-epoxy-4-oxo-7, 10-dodecadienoylamide, irreversibly inactivates yeast fatty acid synthetase. Of all catalytic activities of the synthetase, only the condensation reaction is inhibited by cerulenin. At 0°C and pH 6.5, the second-order rate constant of k=88 M-1•s-1 was obtained for the inactivation by cerulenin. This value was about 90-times greater than the rate constant for the inactivation of the enzyme by iodoacetamide. The enzyme was protected against the action of cerulenin by prior treatment with acetyl-CoA but not malonyl-CoA. Treatment of the enzyme with iodoacetamide, while impairing the synthetase activity, induced malonyl-CoA decarboxylase activity [Kresze, G.-B., Steber, L., Oesterhelt, D., & Lynen, F. (1977) Eur. J. Biochem. 79, 191-199]. Cerulenin had no effect on the malonyl-CoA decarboxylase activity of the iodoacetamide-treated enzyme. N-Ethylmaleimide, in contrast, inhibited the iodoacetamide-induced malonyl-CoA decarboxylase activity. When the enzyme was preincubated with cerulenin, malonyl-CoA decarboxylase activity could not be detected even after treatment of the enzyme with iodoacetamide. These results indicated that the reaction of cerulenin with the peripheral SH-groups of the synthetase is responsible for the inactivation.抄録全体を表示PDF形式でダウンロード (480K) -
Yoshimi YAMAMOTO, Michio FUJIE, Kenji NISHIMURA1982 年 92 巻 1 号 p. 13-21
発行日: 1982年
公開日: 2008/11/18
ジャーナル フリー1) Two forms of acid β-galactosidase [EC 3. 1. 23] with different molecular weights catalyzing the hydrolysis of GM1-ganglioside and p-nitrophenyl-β-D-galactoside were separated and purified from porcine spleen. 2) The apparent molecular weights were 400, 000-600, 000 and 70, 000-74, 000 for the high (termed Am form) and low (termed A1 form) molecular weight forms, respectively. 3) On examination by sodium dodecyl sulfate (SDS)/polyacrylamide gel electrophoresis, both forms of the enzyme had a common protein band of molecular weight 63, 000, and the Am form showed three additional protein bands with molecular weights of 31, 000, 21, 000, and 20, 000. 4) Both forms of the enzyme had similar catalytic functions with regard to pH-optimum, Km, substrate specificity and sensitivity to substrate analogues and other substances such as detergents, bovine serum albumin (BSA) and NaCl. 5) Both forms of the enzyme were fairly stable upon preincubation at 45°C at acidic pH (pH 4.5), but lost their activities at neutral pH (pH 7.0). 6) The A1 form was a monomer at neutral pH (pH 7.0) and formed a dimer at acidic pH (pH 4.5). However, most of the Am form could not be converted to a dimeric form on gel filtration at acidic pH.抄録全体を表示PDF形式でダウンロード (1173K) -
Kei-ichi KAMISANGO, Ikuo SAIKI, Yoshiro TANIO, Hiroyuki OKUMURA, Yoshi ...1982 年 92 巻 1 号 p. 23-33
発行日: 1982年
公開日: 2008/11/18
ジャーナル フリーThe cell-wall skeletons of Listeria monocytogenes strain EGD and Propionibacterium acnes strain C7, which have the ability to induce macrophage activation, were analyzed, and the structures of the peptidoglycans were investigated. The analytical data indicate that both peptidoglycans have glucosamine residues with free amino groups, which are responsible for the resistance to lysozyme. Possible structures of these peptidoglycans were deduced from the composition and the results of determination of N- and C-terminal amino acids, together with the characterization of fragments obtained by enzymatic treatment and partial acid hydrolysis of both peptidoglycans. The results suggested that the peptidoglycan of L. monocytogenes contains a cross-linkage region of peptide chains with meso-diaminopimelic acid and D-alanine, which belongs to the A1γ type (Schleifer, K. H. & Kandler, O. (1972) Bacteriol. Rev. 36, 407-477), whereas the peptidoglycan of P. acnes contains a crosslinkage region of peptide chains with L, L-diaminopimelic acid and D-alanine, in which two glycine residues combine with amino and carboxyl groups of two L, Ldiaminopimelic acid residues. The latter type should be classified as a new type. These cell-wall skeletons and peptidoglycans were shown to have immunoadjuvant activity on the induction of delayed-type hypersensitivity and suppressive activity on the growth of 3-methylcholanthrene-induced fibrosarcoma in BALB/c mice, and the peptidoglycans were shown to be an immunological-active principle of these cell-wall skeletons.抄録全体を表示PDF形式でダウンロード (815K) -
Toshiharu YAGI, Hiroyuki KAGAMIYAMA, Mitsuhiro NOZAKI1982 年 92 巻 1 号 p. 35-43
発行日: 1982年
公開日: 2008/11/18
ジャーナル フリーAspartate: 2-oxoglutarate aminotransferase [EC 2. 6. 1. 1] was purified and crystallized from bakers' yeast. The crystalline preparation gave a single band on polyacrylamide disc gel electrophoresis in the presence of sodium dodecyl sulfate. However, in the absence of sodium dodecyl sulfate, the preparation gave one major band with two faint bands, all of which showed the same specific activity, molecular weight and serological properties. These faint bands appeared to be modified forms produced from the major band during the purification. The enzyme showed a molecular weight of 90, 000±8, 000 and 92, 000±8, 000 by gel filtration and sedimentation equilibrium analysis, respectively. The molecular weight of a subunit was estimated to be 45, 000 by sodium dodecyl sulfate slab gel electrophoresis. Each subunit bound approximately 1 mol of pyridoxal 5'-phosphate. The bound pyridoxal 5'-phosphate showed an absorption maximum at 360 nm (εM: 11, 500) and 430 nm (εM: 8, 200) in alkaline and acidic conditions, respectively. Its protolytic pK was pH 6.3. The enzyme showed an optimum pH of 8.0-9.0, and fairly high amino donor and acceptor specificities; aromatic amino acids and their corresponding 2-oxoacids were catalyzed at rates of 0.2-0.8% of those for aspartate and oxalacetate, respectively. Michaelis constants for various substrate were: L-aspartate (0.11mM), L-glutamate (20.0mM), oxalacetate (0.006mM), and 2-oxoglutarate (0.16mM). The antiserum against yeast aspartate aminotransferase did not form precipitin bands with homogeneous aspartate aminotransferases from pig heart cytosol, pig heart mitochondria or Escherichia coli B.抄録全体を表示PDF形式でダウンロード (2340K) -
Shoji WATABE, Shigeru TAGUCHI, Tadao IKEDA, Minoru TAKADA, Nagasumi YA ...1982 年 92 巻 1 号 p. 45-55
発行日: 1982年
公開日: 2008/11/18
ジャーナル フリーMore than 95% of the apparent cathepsin D activity in the lysosomes of porcine adrenal cortex was due to a genuine cathepsin D that has a molecular weight of 42, 800±800 (mean±standard deviation of the mean in 5 runs) as determined by Sephadex G-100 column chromatography. On CM-Sephadex C-50 column chromatography at pH 6.9, the enzyme was resolved into five peaks which were termed Fractions D1 through D5 in order of their elution from the column. Fraction D4 and Fraction D5 together constituted more than 70% of the total cathepsin D, and were purified through chromatographic procedures to constant specific activities. The content of lysosomal cathepsin D was estimated to be about 1% of the total cellular protein. On isoelectric focusing, Fraction D4 was resolved into one major band with pI of 7.34 (termed Form D4-1) and one minor band of 7.20 (Form D4-2), while Fraction D5 gave one major band with pI of 7.50 (Form D5-1) and two minor ones of 7.37 (Form D5-2) and of 7.20 (Form D5-3). All of these 5 bands were enzymatically active. On SDS-polyacrylamide gel electrophoresis, both Form D4-1 and Form D5-1 dissociated into three subunits with molecular weights of 27, 400±500 (termed Subunit A), 24, 600±400 (Subunit B) and 13, 800±600 (Subunit C) (mean±standard deviation of the mean in 16 determinations). The three subunits all contained carbohydrates.抄録全体を表示PDF形式でダウンロード (1550K) -
Yoshio IMAI1982 年 92 巻 1 号 p. 57-66
発行日: 1982年
公開日: 2008/11/18
ジャーナル フリーInteraction of various polycyclic aromatic hydrocarbons with P-4481 from rabbit liver microsomes was demonstrated by measuring absorption, CD and fluorescence spectra of the hydrocarbons, the cytochrome and the complexes of both. On binding of hydrocarbons such as pyrene and benz[a]anthracene to P-4481, prominent CD peaks appeared at around the wavelengths where the hydrocarbons possess absorption bands. Correspondingly, fluorescence of the hydrocarbons was quenched to various extents depending on the hydrocarbon examined but was restored when the protein structure was destroyed by denaturation. The absorption peaks of the bound hydrocarbons also shifted toward a longer wavelength. Such an appearance of CD bands, quenching of fluorescence emission or a shift of absorption peaks was not seen when the hydrocarbons were mixed with P-4501 or albumin. Titration experiments indicated that hydrocarbons can bind specifically and tightly to P-4481 at a single binding site to form equimolar complexes. Complexes of 21 kinds of hydrocarbons, possessing 2 to 5 benzene rings spread in various directions, were isolated and their spectral properties were investigated. It was shown that the hydrocarbons bind to P-4481 at the same site and compete with one another. Evidence was obtained that the hydrocarbons were bound at the substrate site in the hemecontaining domain of the monooxygenase, P-4481, and metabolized by the aid of NADPH-cytochrome P-450 reductase in the presence of NADPH and O2.抄録全体を表示PDF形式でダウンロード (750K) -
II. Flash Photolysis Studies on Various Hydrocarbon-Bound P-4481 CO CompoundsYoshio IMAI, Tetsutaro IIZUKA, Yuzuru ISHIMURA1982 年 92 巻 1 号 p. 67-75
発行日: 1982年
公開日: 2008/11/18
ジャーナル フリーEffects of aromatic hydrocarbon binding on the photodissociability of CO from ferrous carbonmonoxy P-4481 and their subsequent recombination reactions were studied with the aid of the flash photolysis technique. We found that both size and structure of the bound hydrocarbon are sensitively reflected in the flash photolysis parameters.
The CO aduct of hydrocarbon-free P-4481 was relatively insensitive to light giving a quantum yield of approximately 0.1. The binding of a relatively small hydrocarbon with 2 to 3 fused benzene rings, such as naphthalene and anthracene, had no influence on the photosensitivity, while additions of a larger molecule such as benz[a]anthracene and dibenzanthracenes resulted in a considerable increase in the quantum yield. Such a trend is also seen with the binding of benzoflavone derivatives but not with terphenyls. Among 21 hydrocarbons examined, 7, 8-benzoflavone was most effective in increasing the photodissociability giving a quantum yield of near 1.0.
When the effects of hydrocarbons on the rate of CO recombination were examined, they could be classified into the following three groups. (1) Tricyclic hydrocarbons such as phenanthrene and anthracene, and one of the tetracyclic hydrocarbons, benzo[c]phenanthrene, which enhanced the rate over ten times. (2) Pentacyclic hydrocarbons and their structural analogues such as dibenzanthracenes, 3-methylcholanthrene and 7, 8-benzoflavone, which decreased the rate to below one tenth. (3) Other hydrocarbons of various structures, which did not affect the rate significantly. In the case of pyrene and triphenylene complexes of which photodissociation was not detectable, the CO binding rate was 3-5 times higher than that of the hydrocarbon-free form as judged by the stopped flow method.
Based on these findings, the relationship between the effects and the structures of hydrocarbon molecules was discussed.抄録全体を表示PDF形式でダウンロード (704K) -
III. Effects of Hydrocarbon Binding on the Interaction of Some Ligands with P-4481 HemeYoshio IMAI1982 年 92 巻 1 号 p. 77-88
発行日: 1982年
公開日: 2008/11/18
ジャーナル フリーThe binding of polycyclic hydrocarbons to P-4481 affected the interaction of exogenous ligands, such as ethyl isocyanide and CO in the ferrous state and 1-methyl imidazole in the ferric state, with the heme to various extents depending on the structure of the hydrocarbons. The effect of the hydrocarbons on dissociation constants (Kd) on the three ligands was essentially similar. Kd of ethyl isocyanide and CO for hydrocarbon-free P-4481 were 18 μM and 1.1 μM, respectively. The affinities of both the ligands increased about 3-fold when hydrocarbons of a small molecular size, such as phenanthrene, were bound to P-4481. Addition of a benzene ring to the outside of the phenanthrene molecule resulted in a remarkable decrease in the affinities of both the ligands. The binding of larger hydrocarbons in size produced a stronger inhibitory effect on the interaction of ethyl isocyanide with P-4481. Thus, Kd of the isocyanide for benz[a]anthracene-bound and dibenz[a, c]-anthracene-bound P-4481 were estimated to be 260 μM and 3 mM, respectively. Spectral changes hardly occurred on addition of 4 mM ethyl isocyanide to 3-methylcholanthrene-bound or 7, 8-benzoflavone-bound P-4481. The effect of the hydrocarbon binding on Kd of the ligands was depressed when a benzene ring which links two aromatic rings in the hydrocarbon molecule was replaced by a single C-C bond. A probable structure of the active area of P-4481 composed of the substrate site and the heme is proposed based on the effect of the hydrocarbon binding on the interaction between the exogenous ligands and the P-4481 heme iron reported in this series of papers.
The binding of hydrocarbons to P-4481 increased the intensity ratio of the 430 nm to the 453 nm peak of the absorption spectrum of the ethyl isocyanide compound. When a large and long hydrocarbon in shape such as dibenz[a, h]anthracene was bound to P-4481, a red shift of the Soret absorption peak and modification of the Soret CD pattern of the CO compound were observed.抄録全体を表示PDF形式でダウンロード (871K) -
Ichiro MIYOSHI, Yoichi FUJII, Masaharu NAIKI1982 年 92 巻 1 号 p. 89-94
発行日: 1982年
公開日: 2008/11/18
ジャーナル フリーWe attempted to produce specific antibodies to various gangliosides by immunizing chickens. Antibody activity was determined by enzyme immunoassay (EIA). The optimal conditions for EIA were examined by using chicken anti-GM1 ganglioside (GM1) serum and the final procedure was as follows. Fifty μl of ethanol solution containing 2.5 μg of the glycolipid antigen and 50 μg of taurodeoxycholate were added to each well of an EIA microtitration plate and dried at 37°C for 2 h. Nonspecific sites were blocked by incubation with 1% gelatin-containing buffer, then titration of the chicken antisera was carried out in the antigen-coated plate by incubation at 4°C for 12 h. Next, alkaline phosphatase-labeled anti-chicken IgG (specific antibody, 15 ng; enzyme, 0.054 units) was allowed to react at 37°C for 2 h. The enzyme activity which was bound to the plate was assayed with p-nitrophenol phosphate as substrate.
Under the above conditions, anti-GM1 serum reacted strongly with GM1 and asialo GM1, and anti-GM serum indicated a considerable specificity for GM2. However, we failed to elicit any antibody to GD1a, GD1b, and GT1b. Anti-NeuAcsialosylparagloboside and anti-NeuGc-sialosylparagloboside sera showed a high specificity for the homologous ganglioside. However, anti-NeuGc-hematoside serum reacted equally with both the homologous antigen and NeuGc-sialosylparagloboside, and anti-NeuAc-hematoside (GM3) serum cross-reacted with both N-acetyl and N-glycolyl types of hematoside and sialosylparagloboside.抄録全体を表示PDF形式でダウンロード (451K) -
Akinori KIMURA, Kazunori SHIMADA, Yasuyuki TAKAGI1982 年 92 巻 1 号 p. 95-103
発行日: 1982年
公開日: 2008/11/18
ジャーナル フリーStarting from pAO3, a plasmid consisting of a quarter of colicinogenic factor E1 (ColE1) DNA, various small ColE1 derivatives were constructed by in vitro recombination and their ability to achieve autonomous replication was examined. The 436 base pair HaeIII-C fragment of pAO3 contained information for replication when it was recombined with the non-replicating Amp fragment. However, when it was connected to other DNA fragments, the resulting hybrid molecules were not isolated as plasmids. The present results indicate that the additional region of about 240 base pairs next to the HaeIII-C fragment of ColE1 is also essential for the maintenance of a plasmid state.
Moreover, using various small ColE1 derivatives, the DNA region responsible for the interference and incompatibility functions of ColE1 DNAs was located. The results indicate that the interference and incompatibility functions are coded by the same ColE1 DNA segment and are not essential for the maintenance of a plasmid state.抄録全体を表示PDF形式でダウンロード (2038K) -
Masaaki KODAMA, Takehiko OGATA, Yasuhisa TAKAHASHI, Toshio NIWA, Fumio ...1982 年 92 巻 1 号 p. 105-109
発行日: 1982年
公開日: 2008/11/18
ジャーナル フリーA nontoxic high molecular substance associated with some paralytic shellfish poisons was separated by Sephadex G-50 gel filtration from the toxic digestive glands of the scallop Patinopecten yessoensis fed the causative plankton Protogonyaulax tamarensis. Unlike the corresponding fraction from the nontoxic digestive glands, the substance released gonyautoxins II and III on digestion with RNase T2, suggesting that it is associated with an RNA of P. tamarensis. It is possible that the toxification of scallop is partly due to the toxins already accumulated in Protogonyaulax cells, and partly due to incorporation of this precursor, which releases the toxins as a result of enzymic processes in the shellfish.抄録全体を表示PDF形式でダウンロード (338K) -
Tsukasa SUGANO, Masakazu SHIOTA, Hiroaki KHONO, Masakazu SHIMADA1982 年 92 巻 1 号 p. 111-120
発行日: 1982年
公開日: 2008/11/18
ジャーナル フリーThe effects of glucagon and norepinephrine on gluconeogenesis by perfused chicken liver were studied with fluorimetric monitoring of the redox states of the intracellular pyridine nucleotides.
Glucagon stimulated glucose production from precursors entering the pathway both above and below the level of triose phosphates without causing a detectable effect on the redox states of pyridine nucleotides. Glucagon stimulation was not abolished by subsequent infusion of octanoate or ethanol. The presence of a pyruvate/lactate mixture plus NH4C1 resulted in a maximum efficacy of glucagon. Glucose production from lactate and fructose was stimulated by norepinephrine. Norepinephrine always caused a change towards increased reduction of pyridine nucleotides with an increase in the β-hydroxybutyrate/acetoacetate ratio, but displayed no stimulation of glucose and lactate production from pyruvate. As a result of octanoate infusion with lactate, the changes induced by norepinephrine were reversed. The responses to norepinephrine and phenylephrine were decreased markedly in liver perfused with a calcium-free medium and/or with phentolamine. Infusion of calcium into the calcium-deficient liver caused an abrupt elevation of glucose production together with a reduction of pyridine nucleotides, and the original response to norepinephrine was recovered.
The results demonstrate that the effects of glucagon and norepinephrine on gluconeogenesis are not identical, and that norepinephrine stimulation is mediated through an α-adrenergic and calcium-dependent mechanism in which redox changes of mitochondrial pyridine nucleotides are involved.抄録全体を表示PDF形式でダウンロード (792K) -
Satoru KUHARA, Emiko EZAKI, Tamo FUKAMIZO, Katsuya HAYASHI1982 年 92 巻 1 号 p. 121-127
発行日: 1982年
公開日: 2008/11/18
ジャーナル フリーThe binding constant and binding free energy of each subsite of lysozyme upon substrate binding have been customarily estimated from the experimental data with assumptions regarding the binding mode of substrate3 and the additivity of binding free energy of each subsite. In the present study, the binding constants and binding free energy of subsites were estimated from experimentally obtained overall binding constants on native and Trp 62-modified lysozymes. The estimations of binding constants and binding free energy were carried out by an optimization method, the modified Powell method, without assuming the binding mode for substrate. First the binding free energies of subsites A, B, and C were estimated from the experimental binding constants of (GlcNAc)1 to (GlcNAc)3, and the binding free energies of subsites D, E, and F were determined from the estimated free energies of subsites A, B, and C, and the experimentally obtained reaction time-courses of substrate (GlcNAc)5. Finally, the values of three rate constants in the lysozyme-catalyzed reaction of chitooligosaccharide were estimated from the experimental time-course by using the binding free energies obtained by the modified Powell method.抄録全体を表示PDF形式でダウンロード (564K) -
Yasutsugu FUKUNAGA, Yasuhiro KATSURAGI, Takashi IZUMI, Fumio SAKIYAMA1982 年 92 巻 1 号 p. 129-141
発行日: 1982年
公開日: 2008/11/18
ジャーナル フリーSeveral kynurenine derivatives including N'-formylkynurenine were prepared in high purity by the ozonization of the corresponding indole compounds. The fluorescence characteristics of those derivatives were examined in connection with the use of their fluorophores as reporters for the local environment of tryptophan in proteins.
Kynurenine is a weak emitter of fluorescence, with an emission maximum at 480 nm on excitation at 365 nm. With decreasing solvent polarity, the fluorescence intensity increases logarithmically and the emission maximum shifts to blue. A linear relation between these fluorescence characteristics and solvent polarity exists when the polarity is shown in terms of dielectric constant.
N'-Formylkynurenine is a somewhat stronger emitter of fluorescence than kynurenine. The emission maximum is 434 nm on excitation at 325 nm and it shifts to blue in solvents of low polarity. This blue shift is also linear with respect to the dielectric constant of the solvent.
Other factors influencing kynurenine fluorescence and N'-formylkynurenine fluorescence examined were neighboring groups, ionic strength, temperature, and protein denaturants.
Based on the results of the present investigation, the local environment of tryptophan 62 in hen egg-white lysozyme was examined using Kyn 62-lysozyme.抄録全体を表示PDF形式でダウンロード (1040K) -
Yasutsugu FUKUNAGA, Hidetsune TAMAOKI, Fumio SAKIYAMA, Kozo NARITA1982 年 92 巻 1 号 p. 143-153
発行日: 1982年
公開日: 2008/11/18
ジャーナル フリーThe previously reported method for the preparation of Kyn 59-RNase T1 and NFK 59-RNase T1 has been improved, and these two proteins have been obtained in high purity. Kyn 59-RNase T1, fully active for the hydrolysis of GpA and GpC, emitted a 35-fold-enhanced fluorescence of kynurenine relative to acetylkynurenine amide with an emission maximum at 455 nm upon excitation at 380 nm. The polarity of the environment of Kyn 59 estimated from the emission maximum corresponded to a dielectric constant of 6. Upon excitation at 325 nm, NFK 59-RNase T1, less active than Kyn 59-RNase T1, exhibited a quenched N'-formylkynurenine fluorescence with an emission maximum at 423 nm, from which the value of 12 was obtained as the dielectric constant of the surroundings of residue 59.
In both modified proteins, distinct tyrosine fluorescence appeared on excitation at 280 nm. The detection of an energy transfer from tyrosine to residue 59 suggests that the tertiary structure is very similar in Kyn 59-RNase T1 and native RNase T1.
With guanidine hydrochloride, Kyn 59-RNase T1 was less stable than the native protein. Carboxymethylation at Glu 58 was shown to stabilize the active site of the modified enzyme.
Based on the information collected for Kyn 59-RNase T1, the local environment and possible roles of the sole tryptophan residue in RNase T1 are discussed.抄録全体を表示PDF形式でダウンロード (857K) -
Yasutsugu FUKUNAGA, Fumio SAKIYAMA1982 年 92 巻 1 号 p. 155-161
発行日: 1982年
公開日: 2008/11/18
ジャーナル フリーFluorescence titrations of kynurenine and tyrosine in Kyn 59-RNase T1 and NFK 59-RNase T1 were carried out by monitoring protein fluorescence through a pH change from 1.5 to 10.5.
In the titration of kynurenine fluorescence at 455 nm, a few small but distinct quenching events occurred between pH 3.5 and 9.5. Three ionizable groups were found to be responsible for the individual steps of quenching observed. These groups are Glu 58 with pKa 4.6, His 40 or 92 with pKa 7.8 and Lys 41 with pKa 8.7. From this result, a subtle conformational change associated with the proton dissociation equilibria of Glu 58 and His 40 or 92 in the active site of Kyn 59-RNase T1 is suggested. The pH-titration behavior of tyrosine fluorescence in Kyn 59-RNase T1 was different from that of kynurenine fluorescence. Two acidic groups with pKa's 3.2 and 6.5 were detected as perturbants.
In NFK 59-RNase T1, both N'-formylkynurenine and tyrosine showed almost the same fluorescence behavior during titration, which was characterized by two transitions between pH 3 and 8 in each titration curve. Two ionizable groups with pKa's 3.7-3.8 and 6.7-6.8 were determined. The role of the latter ionizable group is discussed in relation to the enzyme function of RNase T1.
From the close similarity in structure and function between Kyn 59-RNase T1 and RNase T1, it is suggested that the same mechanism of conformational change linked to the ionization states of Glu 58 and His 40 or 92 exists in the native protein too.抄録全体を表示PDF形式でダウンロード (565K) -
Seiko SHIGETA, Akira MATSUDA, Satoru OKA1982 年 92 巻 1 号 p. 163-172
発行日: 1982年
公開日: 2008/11/18
ジャーナル フリーA β-N-acetylhexosaminidase [EC 3.2.1.30] was isolated from internal organs of the sea-squirt, Styela plicata. The enzyme was purified 1, 560-fold in 5% yield. The preparation was fairly homogeneous as examined by disc and SDS polyacrylamide gel electrophoresis and Sephadex G-200 chromatography. The molecular weight of the enzyme was estimated to be 132, 000 by gel chromatography and 66, 000 by SDS polyacrylamide gel electrophoresis. Therefore, this β-N-acetylhexosaminidase was considered to be a dimer. The optimum pH for activity was 4.0 but the enzyme was stable in the pH range from 5 to 6. The isoelectric point was 4.99. This enzyme was inhibited by Fe2+ Hg2+, Ag+, and PCMB but not by acetate. The isolated enzyme hydrolyzed both p-nitrophenyl-N-acetyl-β-D-glucosaminide and p-nitrophenyl-N-acetyl-β-D-galactosaminide. The hydrolysis rate of p-nitrophenyl-N-acetyl-β-D-galactosaminide was 43% of that of p-nitrophenyl-N-acetyl-β-D-glucosaminide. The enzyme liberated N-acetylhexosamine from asialodegalactosyl ovomucoid glycopeptide, asialodegalactosyl fetuin glycopeptide and the fragment of hyaluronic acid prepared by hyaluronidase treatment.抄録全体を表示PDF形式でダウンロード (1181K) -
II. Interaction with Guanine Nucleotides and Aminoacyl-tRNASeisuke HATTORI, Kentaro IWASAKI1982 年 92 巻 1 号 p. 173-183
発行日: 1982年
公開日: 2008/11/18
ジャーナル フリーThe high molecular weight form of polypeptide chain elongation factor-1, or EF-1H, has been purified from pig liver to apparent homogeneity and shown to be a 1:1:1 complex of EF-1α, EF-1β, and EF-1γ, namely, EF-1αβγ (Hattori, S. and Iwasaki, K. (1980) J. Biochem. 88, 725-736). Therefore, its enzymatic properties were investigated in detail. The effects of various components such as NH4Cl, Mg(CH3COO)2, guanine nucleotides and Phe-tRNA on its heat stability were investigated and it was found that its properties were very close to those of the aminoacyl-tRNA binding enzyme from rabbit reticulocytes (McKeehan, W. L. and Hardesty, B. (1969) J. Biol. Chem. 244, 4330-4339). Furthermore, it was reported that EF-1αβγ seemed to dissociate into EF-1α and EF-1βγ when guanine nucleotides were present, and also to form a complex of EF-1α•GTP•[14C]Phe-tRNA when both GTP and [14C]PhetRNA were present. On the contrary, however, when the interaction between EF-αβγ and guanine nucleotides was analyzed by gel filtration at 4°C, the dissociation of EF-1αβγ could not be detected, instead the formation of binary complexes containing EF-1αβγ and guanine nucleotides was observed. The dissociation constants of the EF-1αβγ•GDP and EF-1αβγ•GDP complexes were estimated to be 6.8×10-6M and 7.3×10-6M, respectively, and the number of binding sites per molecule of EF-1αβγ for these nucleotides was estimated to be one. Little, if any, interaction was detected between EF-11αβγ and aminoacyl-tRNA even in the presence of GTP. The controversial results obtained by different methods as described above seemed to be compatible with each other, if one assumes that the dissociation constant for the conversion of EF-1αβγ into EF-1α a and EF-1βγ increased with the increase in temperature. When gel filtration of EF-1αβγ was carried out with a solution containing both GTP and [14C]Phe-tRNA, the formation of a ternary complex containing EF-1α, GTP and [14C]Phe-tRNA was detected at 4°C, although the amount was very small. From these results, we assumed that EF-1αβγ interacts with GTP and aminoacyl-tRNA (aa-tRNA) according to the following sequences;抄録全体を表示PDF形式でダウンロード (966K) -
Hideo SAWADA, Akira HARA, Toshihiro NAKAYAMA, Masakazu HAYASHIBARA1982 年 92 巻 1 号 p. 185-191
発行日: 1982年
公開日: 2008/11/18
ジャーナル フリーThe kinetic properties of the NADPH-dependent reduction of aromatic aldehydes and ketones catalyzed by low- and high-molecular-weight aldehyde reductases [alcohol:NADP oxidoreductase, EC 1.1.1.2] and 3α- and 3β-hydroxysteroid dehydrogenases [EC 1.1.1.50 and 1.1.1.51] of rabbit liver were compared. Initial velocity measurements with pyridine-4-aldehyde, 4-benzoylpyridine and androstadione as substrates and inhibition studies with their products indicated that all the enzymes followed an ordered Bi Bi reaction mechanism with coenzyme binding first and leaving last. However, phenylpyruvic acid inhibited 3α-hydroxysteroid dehydrogenase and low-molecular-weight aldehyde reductase noncompetitively with respect to either NADPH or substrate, whereas it inhibited 3β-hydroxysteroid dehydrogenase and high-molecular-weight aldehyde reductase uncompetitively. Cibacron blue F3GA dye was a dead-end inhibitor of the enzymes, being competitive with respect to NADPH and noncompetitive with respect to the other substrate, but the K1 value of 3α-hydroxysteroid dehydrogenase for this dye was much higher than those of the other enzymes.抄録全体を表示PDF形式でダウンロード (506K) -
I. Ouabain-Sensitive Alternative Binding of Three Na+ or Two K+ to the EnzymeHideo MATSUI, Haruo HOMAREDA1982 年 92 巻 1 号 p. 193-217
発行日: 1982年
公開日: 2008/11/18
ジャーナル フリーBy a centrifugation method with purified Na+, K+-ATPase and 22Na+ or 42K+, the specific binding of Na+ or K+ to the enzyme was determined as ouabain-sensitive binding, the difference between that in the absence and presence of 0.1mM ouabain. The ouabain-sensitive K+ binding at 50 μm KCl was 5.7 nmol/mg protein ([3H]-ouabain binding=2.8 nmol/mg protein). It was completely inhibited by 10mM NaCl but not affected by 10mM choline chloride. The binding was partially inhibited by 0.2mM ATP and by 5mM MgCl2 but not by 0.2mM P1. At KCl concentrations of more than 20 μm the ouabain-sensitive K+ binding reached a saturation level twice as high as that of ouabain binding. The apparent Kd for K+ was 6 μM. A Scatchard plot of K+ binding showed a curved line, suggesting positive cooperativity of two binding sites for K+ (Hill coefficient nH=1.72); the two Kd's were estimated to be 36 μM and 1 μM.
The ouabain-sensitive Na+ binding at 0.5 mM NaCl was 5.3 nmol/mg protein (ouabain binding=2.5 nmol/mg protein). It was specifically inhibited by 0.1mM KCl but not affected by 10mM choline chloride. The Na+ binding was inhibited by 5mM MgCl2 but not by 0.2mM ATP or by 0.2mM P1, contrasting with the K+ binding. Oligomycin increased the Na+ binding to 6.4 nmol/mg protein. This stimulation was much more pronounced at 0.1mM NaCl; from 1.6 nmol to 6.1 nmol Na+/mg protein. Without oligomycin the Na+ binding at NaCl concentrations of less than 1mM did not reach saturation because of the enzyme's low affinity for Nat With oligomycin the binding at NaCl concentrations of more than 0.2 mM reached a saturation level which was 2.7-fold that of ouabain binding. The apparent Kd's for Na+ in the absence and presence of oligomycin were about 340 μm and 60 μm respectively. The affinity enhancement by oligomycin appears to be due to an increase in cooperativity between the three binding sites for Na+ (nH=1.23→1.54).
Simultaneous measurement of Na+ and K+ binding at a constant Na+ concentration of 0.5mM or 0.1 mM with oligomycin and increasing concentrations of K+ showed a decreasing pattern of ouabain-sensitive Na+ binding which was mirrored exactly by an increasing pattern of ouabain-sensitive K+ binding. It is concluded that 3Na+ and 2K+ do not bind simultaneously to the same enzyme molecule but alternatively to the different conformers of the enzyme, namely E1 or E2.抄録全体を表示PDF形式でダウンロード (2178K) -
II. General Properties of Ouabain-Sensitive K+ BindingHaruo HOMAREDA, Hideo MATSUI1982 年 92 巻 1 号 p. 219-231
発行日: 1982年
公開日: 2008/11/18
ジャーナル フリーGeneral properties of ouabain-sensitive K+ binding to purified Na+, K+-ATPase [EC 3.6.1.3] were studied by a centrifugation method with 42K+. 1) The affinity for K+ was constant at pH values higher than 6.4, and decreased at pH values lower than 6.4. 2) Mg2+ competitively inhibited the K+ binding. The dissociation constant (Kd) for Mg2+ of the enzyme was estimated to be about 1mM, and the ratio of Kd for Mg+ to Kd for K+ was 120:1. The order of inhibitory efficiency of divalent cations toward the K+ binding was Ba2+_??_Ca2+>Zn2+_??_Mn2+Sr2+>Co2+>Ni2+>Mg2+. 3) The order of displacement efficiency of monovalent cations toward the K+ binding in the presence or absence of Mg2+ was Tl+> Rb+≥(K+)>NH4+≥Cs+>Na+>Li+. The inhibition patterns of Na+ and Li+ were different from those of other monovalent cations, which competitively inhibited the K+ binding. 4) The K+ binding was not influenced by different anions, such as Cl-, SO2-4, NO-3, acetate, and glycylglycine, which were used for preparing imidazole buffers. 5) Gramicidin D and valinomycin did not affect the K+ binding, though the former (10 μg/ml) inhibited the Na+, K+-ATPase activity by about half. Among various inhibitors of the ATPase, 0.1mM p-chloromercuribenzoate and 0.1mM tri-n-butyltin chloride completely inhibited the K+ binding. Oligomycin (10 μg/gimp and 10mM N-ethylmaleimide had no effect on the K+ binding. In the presence of Na+, however, oligomycin decreased the K+ binding by increasing the inhibitory effect of Na+, whether Mg2+ was present or not. 6) ATP, adenylylimido diphosphate and ADP each at 0.2 mM decreased the K+ binding to about one-fourth of the original level at 10 μm K+ without MgCl2 and at 60 μm K+ with 5 mM MgCl2. On the other hand, AMP, P1, and p-nitrophenylphosphate each at 0.2mM had little effect on the binding.抄録全体を表示PDF形式でダウンロード (1084K) -
Hazime SAITO, Mayumi KAMEYAMA, Masahiko KODAMA, Chikayoshi NAGATA1982 年 92 巻 1 号 p. 233-241
発行日: 1982年
公開日: 2008/11/18
ジャーナル フリーA 13C NMR study of calf thymus histone H1 in both aqueous solution and 2-chloroethanol solution was performed to clarify the folding behavior in these systems. To ascertain the general trend of displacements of 13C shifts upon folding in an enhanced manner, the latter solvent was employed since it is known to increase the amount of α-helix content in histone to about 50%. Generally, upfield displacements of Cβ signals (up to 1.4 ppm) were clearly identified as helix-induced peaks, although displacements of Cα signals, which might be much larger, were not easily distinguished because of overlap of several broadened signals with reduced peak intensities. In particular, we found that the upfield displacement of Ala Cβ, by 1.1 ppm, is an excellent probe to monitor the presence of α-helix conformation in both 2-chloroethanol and aqueous solutions. This upfield displacement of the Cβ signal in α-helix segment is consistent with our previous findings for a number of model polypeptides by ordinary and solid-state high resolution 13C NMR spectroscopy. Further, we observed that 13C peaks of several residues (Tyr, Ser, Leu, Ile, and Val) were suppressed as a result of specific folding of H1 in the presence of NaCl in aqueous solution. Thus, it appears that several tightly-folded segments whose 13C signals were considerably broadened are located in the central core portion.抄録全体を表示PDF形式でダウンロード (641K) -
Hiroshi KIMURA, Yoshitaka FUTAMI, Sei-ichiro TARUI, Takaaki SHINOMIYA1982 年 92 巻 1 号 p. 243-251
発行日: 1982年
公開日: 2008/11/18
ジャーナル フリーWe examined the effects of calcium, bile salts and colipase, a protein cofactor, on pancreatic lipase purified from human duodenal juice. It was found that 10% gum acacia, used as a stabilizer of triglyceride emulsions, was contaminated with a high concentration of calcium-between 14 and 20mM. Olive oil stabilized with gelatin was used as a substrate to decrease the calcium content in the reaction mixture. The activity of duodenal juice and pancreatic extracts was inhibited by 50% with 10 μm EGTA. Ca2+ activated pancreatic lipase, irrespective of the presence of bile salts plus colipase. The apparent Ka for Ca2+ was calculated to be 10μM and this was not affected by colipase and bile salts. However, maximal activation by Ca2+ required both bile salts and colipase. Ca2+ increased Vmax of the reaction and decreased the apparent Km for the substrate in the presence of both colipase and bile salts. However, no binding of 45Ca to colipase was demonstrated. High concentrations of bile salts inhibited the activity in the presence of either colipase or Ca2+, while in the presence of both colipase and Ca2+ a marked activation by bile salts was observed. Colipase by itself also activated lipase.
We conclude from the present experiments that the activity of human pancreatic lipase is regulated by three effectors; Ca2+, bile salts and colipase, which may enhance each other's efficiency in the enzyme activation. In addition, we suggest that an activation rather than an inhibition of pancreatic lipase by bile salts is more significant physiologically, because it is well-known that bile salts play an important role in fat absorption in the intestine.抄録全体を表示PDF形式でダウンロード (717K) -
Masataka ISHINAGA, Junichi SATO, Yasuo KITAGAWA, Etsuro SUGIMOTO, Mako ...1982 年 92 巻 1 号 p. 253-263
発行日: 1982年
公開日: 2008/11/18
ジャーナル フリーErucic acid was incorporated into cardiac phosphatidylserine and hepatic and cardiac sphingomyelin of male Sprague-Dawley rat. The fatty acid compositions of mitochondrial and microsomal phospholipids were similar in both liver and heart. The effect of a low fat diet and a diet containing erucic acid on the fatty acid composition of mitochondrial and microsomal phospholipids was also similar, except for the effect on sphingomyelin. However, the diet containing erucic acid influenced the metabolism of phosphatidylcholine of the heart but not of the liver, indicating that the turnover of 1-stearoyl-2-arachidonoyl phosphatidylcholine in the heart was inhibited by the diet containing erucic acid. On the other hand, the proportion of erucic acid in the free fatty acid was higher in the heart than in the liver.抄録全体を表示PDF形式でダウンロード (700K) -
Sumihiro HASE, Ken-ichi FUJIMURA, Makoto KANOH, Tokuji IKENAKA1982 年 92 巻 1 号 p. 265-270
発行日: 1982年
公開日: 2008/11/18
ジャーナル フリーCrystalline Taka-amylase A, prepared from Takadiastase, was fractionated into four fractions by DEAE-Sephacel and Concanavalin A-Sepharose column chromatography. The relative weight ratio of the fractions was 90:4:4:2. They had similar molecular weights (51, 000), amino acid compositions, and hydrolytic activity against soluble starch, but different phenyl maltosidase activities and electrophoretic mobilities on polyacrylamide gel electrophoresis. Three of the fractions mainly had the high mannose type sugar chain with the sugar composition of Man5- GlcNAc2, but the other fraction had only one N-acetylglucosamine residue as the sugar chain. These results suggested that Taka-amylase A was heterogeneous both in the sugar portions and in the polypeptide portions.抄録全体を表示PDF形式でダウンロード (898K) -
Norio MURASE, Kinji GONDA1982 年 92 巻 1 号 p. 271-273
発行日: 1982年
公開日: 2008/11/18
ジャーナル フリーThe interaction between liposomes and clay was investigated. Lecithin liposomes were adsorbed by montmorillonite, and aggregation of the clay particles occurred. From studies using differential scanning calorimetry (DSC), the transition temperature (Tc) from a gel to a liquid-crystalline state of this phospholipid increased by several degrees on adsorption by the clay. Although a lecithin molecule as a whole seems to have no net charge, liposomes were considered to have been adsorbed by the clay via the choline positive charge, which occupies the outermost part of the liposome membrane, as the clay is known to be negatively charged.抄録全体を表示PDF形式でダウンロード (219K) -
Tadashi YOSHIMOTO, Taizo KITA, Makoto ICHINOSE, Daisuke TSURU1982 年 92 巻 1 号 p. 275-282
発行日: 1982年
公開日: 2008/11/18
ジャーナル フリーDipeptidyl aminopeptidase IV [EC 3.4.14.5] was purified from the water extract of porcine pancreas acetone powder by a series of column chromatographies on DEAE-Sephadex and gel filtration on Sephadex G-200, and was finally subjected to gel filtration on Toyo-pearl in the presence of 1% deoxycholate. The purified enzyme was homogeneous as judged by disc gel and SDS gel electrophoreses. The enzyme was most active at pH 8.0 with Gly-Pro-β-naphthylamide (Gly-Pro-2-NNap) as the substrate and hydrolyzed peptide bonds involving the carboxyl group of prolyl residues penultimate to unprotected termini. The enzyme was completely inactivated by diisopropyl phosphorofluoridate (DFP), but only slightly inhibited by phenylmethane sulfonylfluoride (PMSF), SH-blocking reagents and metal chelators. The isoelectric point of the enzyme was 4.8, and the molecular weight was estimated to be 230, 000 by gel filtration on Sephadex G-200 and 115, 000 by sodium dodecyl sulfate (SDS) gel electrophoresis, suggesting that the enzyme is composed of two identical subunits.抄録全体を表示PDF形式でダウンロード (1508K) -
Tsuguo MIZUOCHI, Takahiro TANIGUCHI, Yoshio ASAMI, Junki TAKAMATSU, Ma ...1982 年 92 巻 1 号 p. 283-293
発行日: 1982年
公開日: 2008/11/18
ジャーナル フリーHuman fibrinogen contains four asparagine-linked sugar chains in one molecule. All Bβ and γ subunits obtained from both normal fibrinogen and abnormal fibrinogen Nagoya contain 1 mol each of an asparagine-linked sugar chain. The sugar chains were quantitatively liberated as radioactive oligosaccharides from the polypeptide portion by hydrazinolysis followed by N-acetylation and NaB3H4 reduction. By the combination of sequential exoglycosidase digestion and methylation analysis, the structures of the sugar chains of human fibrinogen were elucidated to be NeuAcα2→6Galβ1→4GleNAcβ1→2Manα1→6(NeuAcα2→ 6Galβ1→4GlcNAcβ1→2Manα1→3)Manβ1→4GIcNAcβ1 →4GlcNAc and Galβ1→4GlcNIAcβ1→2Manα1→6(NeuAcα2→6Galβ1 →4GlcNAcβ1→2Manα1→ 3)Manβ1→4GlcNAcβ1→4GlcNAc. Neither quantitative nor qualitative differences were found between the sugar chain moieties of normal fibrinogen and fibrinogen Nagoya, indicating that the molecular basis of the abnormality in the latter may reside in its polypeptide moieties.抄録全体を表示PDF形式でダウンロード (864K) -
Masashi KOSAKAI, Zensaku YOSIZAWA1982 年 92 巻 1 号 p. 295-303
発行日: 1982年
公開日: 2008/11/18
ジャーナル フリーEleven purified sulfated oligosaccharides previously isolated from the deamination products of porcine and whale heparins were coupled with a fluorescent compound, 2-aminopyridine. The resulting pyridylamino derivatives were used as standards in the development of high performance liquid chromatography procedures for their separation and quantitation on a itbondapak-NK, , anion-exchange column. Four steps of isocratic elution with 30% methanol solution containing 10mM ammonium acetate (pH 5.5), 30% methanol solution containing 20mM ammonium acetate (pH 5.5), 10% methanol solution containing 20mM ammonium acetate and 20mM ammonium phosphate (pH 5.5), and 10% methanol solution containing 20mM ammonium acetate and 30mM ammonium phosphate (pH 5.5) satisfactorily separated the standard pyridylamino derivatives of monosulfated disaccharides (Di-6S(G), Di-6S, and Di-2S), disulfated disaccharide (Di-2, 6S) plus monosulfated tetrasaccharides (Tetra-6'S and Tetra-2S) plus monosulfated trisaccharide (Tri-6'S), disulfated tetrasaccharides (Tetra-6, 6'S and Tetra-2, 6S), and disulfated tetrasaccharides (Tetra-6, 6'S and Tetra-2, 6'S), respectively. The pyridylamino derivatives of the deamination products of porcine and whale heparins were separated into 19 peaks, including 11 peaks corresponding to the standards, by the foregoing procedures. Linear gradient elution with 10% methanol solution containing 10mM ammonium acetate (pH 5.5)-10% methanol solution containing 20mM ammonium acetate and 0.1M ammonium phosphate (pH 5.5) directly separated the pyridylamino derivatives of the deamination products of porcine and whale heparins. The proportions of the pyridylamino-saccharides in the 19 peaks from porcine and whale heparins were then calculated in terms of the absorbance at 309 nm in total. The structures抄録全体を表示PDF形式でダウンロード (1309K) -
Sumio KAWATA, Toshihiro SUGIYAMA, Kouichi SEKI, Seiichiro TARUI, Mitsu ...1982 年 92 巻 1 号 p. 305-313
発行日: 1982年
公開日: 2008/11/18
ジャーナル フリーThe relationship between the changes in the amount of the components of the liver microsomal electron transport systems and drug hydroxylase activities on administration of p-nitroanisole was investigated. The content of cytochrome P-450 in the p-nitroanisole-treated rats was not significantly different from that in the controls during the treatment. The cytochrome b5 content increased 2.2-fold over that of the controls after 14 days of treatment. Demethylation activities per mg microsomal protein of p-nitroanisole, aminopyrine, and benzphetamine were enhanced 2.7-, 2.4-, and 2.8-fold, respectively, but aniline hydroxylase activity was not enhanced. Similar results were obtained with microsomes from diisopropyl 1, 3-dithiol-2-ylidenemalonate (NKK-105)-treated rats. The time-course study of changes in the amount of the components of drug hydroxylase systems suggested that the increase in cytochrome b5 content enhanced the demethylation activities. In the reconstituted system containing cytochrome P-450 partially purified from p-nitroanisole- or NKK-105-treated rats, cytochrome b5 was required for the maximal activities of the demethylation reactions, but did not participate in aniline hydroxylation. These findings showed good correlation between the change of cytochrome b5 content and the demethylation activities and suggested that the increase in cytochrome b5 content might enhance the demethylation activity by stimulating supply of a second electron to cytochrome P-450.抄録全体を表示PDF形式でダウンロード (771K) -
Susumu OHARA, Kaijiro ANZAI, Toshiwo ANDOH1982 年 92 巻 1 号 p. 315-318
発行日: 1982年
公開日: 2008/11/18
ジャーナル フリーWe described methods of preparation of folding proteins of DNA in quantity for further characterization. In principle folding proteins were first isolated as a specific DNA-protein complex from the bulk cellular proteins by centrifugation through a sucrose density gradient, followed by dissociation or by passage through a SH-Sepharose column. The overall yield was 105 μg of folding proteins from 1.5×109 cells. The folding protein preparation was shown to be composed of a mixture of a few kinds of SH-containing proteins, the main components being of molecular weight of 52K and 60K.抄録全体を表示PDF形式でダウンロード (1592K)
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