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1.
Hiroshi Kanazawa Toshiaki Kayano Tatsuya Kiyasu Masamitsu Futai 《Biochemical and biophysical research communications》1982,105(4):1257-1264
The 1855-nucleotide long DNA sequence of part of the gene cluster for the proton-translocating ATPase from was determined by the method of Maxam-Gilbert. The sequence covers the genes for the β and ε subunits of F1 along with the flanking region. The amino acid sequence of these subunits deduced from the nucleotide sequence indicates that the β and ε subunits have 459 and 138 amino acids, respectively. The possible secondary structure of the both subunits was estimated from the deduced primary structures. A possible nucleotide binding site in the β subunit is also discussed on the basis of the primary and secondary structures. The codons used in the genes for all the components of F1F0 were different in different genes, suggesting that the amount of each subunit in the F1F0 is determined to some extent on a translational level. 相似文献
2.
Isolated rat hepatocytes were labeled with [35S]methionine in the absence or presence of cycloheximide or chloramphenicol. The cytochrome 1 complex was isolated from labeled cells by a micromethod and analyzed by SDS-polyacrylamide gel electrophoresis and fluorography. All subunits except the two smallest, subunits VII and VIII, were labeled in the absence of translational inhibitors. In the presence of cycloheximide only subunit III (molecular weight, 30 000) was labeled. This polypeptide, identified as an apo-cytochrome b, was weakly labeled with [35S]methionine in the presence of cycloheximide, indicating a strict dependence of cytoplasmically synthesized products for its assembly. In the presence of chloramphenicol, labeling was inhibited only in subunit III. 相似文献
3.
Nucleotide sequence of the genes for F0 components of the proton-translocating ATPase from Escherichia coli: prediction of the primary structure of F0 subunits 总被引:22,自引:0,他引:22
H Kanazawa K Mabuchi T Kayano T Noumi T Sekiya M Futai 《Biochemical and biophysical research communications》1981,103(2):613-620
The 1763 nucleotide-long-DNA sequence of part of the gene cluster for the proton-translocating ATPase from was determined. The sequence covers the genes for the a and b subunits of F0 along with the intercistronic regions. In the region preceding the gene for the a subunit, a reading frame encompassing 127 amino acids was found. The primary structure of the a and b subunits were deduced and the properties of these proteins were predicted. Analysis of codon usage in these genes was made. 相似文献
4.
C.Channa Reddy Nan-qian Li Chen-Pei D. Tu 《Biochemical and biophysical research communications》1984,121(3):1014-1020
A new glutathione -transferase has been purified to homogeneity from 105,000 × g supernatant of Sprague-Dawley rat liver homogenates. The purified enzyme exhibited specific activities of approximately 1.8, and 0.12 μmoles. min?1. mg?1 toward 1-chloro 2,4-dinitrobenzene and cumene hydroperoxide respectively. The SDS gel electrophoresis data on subunit composition revealed that the new transferase is composed of two subunits with an identical Mr of 24,400 (Yα Family). Our translation experiments with rat liver poly(A) RNAs and substrate specificity data suggest that this subunit is different from the previously reported Ya, Yb and Yc subunits of rat liver glutathione -transferases. Comparatively, the new isozyme showed significant activity toward 1,2 epoxy-3-(P-nitrophenoxy)-propane, ethacrynic acid and P-nitrophenyl acetate, 0.4, 0.34 and 0.18 μ moles. min?1. mg?1 respectively. 相似文献
5.
Potato virus X (PVX) is modified by incubation with chlorogenic acid and polyphenoloxidase. The product made at pH 7 (PVX-Q1) is grey in colour, retains about 2/3 of its initial infectivity, and contains, on average, 1 molecule of bound chlorogenic acid per protein subunit. The product made at pH 7.8 (PVX-Q2) is blue, retains at least of its infectivity, and contains approximately 2 molecules of chlorogenic acid per subunit. Both preparations contain a proportion (18–42%) of cross linked subunits. Brief exposure to trypsin converts subunits of both types of PVX-Q to a form with a slightly lower MW; the reaction goes more extensively with PVX-Q1 (80% converted) than with PVX-Q2 (45% converted). Prolonged exposure to trypsin degrades both forms of PVX-Q to free quinic acid and peptides, apparently only one of which contains chlorogenic acid. It is argued that, in PVX-Q1, predominantly one specific lysine ε-NH2 has been modified with chlorogenoquinone. The structure of PVX-Q2 is less clear. 相似文献
6.
The surface activity and enzymic properties of the factor F1, the catalytic moiety of Streptococcus faecalis H+-ATPase, has been studied at the air-water and phospholipid-water interfaces. F1 does not interact with the monolayer phospholipids, hence its adsorption on a biological membrane must be due mainly to its recognition of proteins of the hydrophobic complex. The dimensions of the F1 molecule at the air-water interface have been estimated. In the presence of Mg2+, base area is , height . Bearing in mind the size of a globular subunit, it follows from the measurements that the major F1 subunits should all lie in the same plane. The ATPase activity of F1 at the interface is inversely proportional to the monolayer density. With low density monolayer, the specific ATPase activity is higher at the interface than in the bulk of the solution.Adsorption of F1 at the interface shifts the isoelectric point of the protein, apparently due to changes in its conformation. The findings are discussed relative to the proton-active transport mechanism. 相似文献
7.
Interchain disulfide bridges in ribonuclease BS-1 总被引:3,自引:0,他引:3
RNAase BS-1, a dimeric ribonuclease isolated from bovine seminal plasma, is made up of two identical subunits whose amino acid sequence is homologous to the sequence of bovine pancreatic RNAase A. The dimeric structure, resistant to denaturating agents, is sensitive to thiol reagents even in the absence of denaturants. The isolation and characterization of a cystine peptide containing two adjacent residues is reported. As the peptide molecular weight is halved after reductive cleavage with dithiothreitol, a structure based on two interchain disulfide bonds between the two adjacent of each subunit is proposed. The singularity of such a structure for a small enzymatic protein is discussed. 相似文献
8.
L Winqvist L Eriksson G Dallner B Ersson 《Biochemical and biophysical research communications》1976,68(3):1020-1026
Four subunits of the acetylcholine receptor molecule, obtained from the electric organ of , have been isolated using polyacrylamide gel electrophoresis, and assayed by titration with a fluorescent lanthanide, terbium, and by affinity-labeling with -(-maleimido)benzyl [trimethyl-3H] ammonium iodide. The site with which the activator-analogue affinity label reacts, as well as the terbium-binding sites, are mainly associated with the smallest of the subunits of an apparent molecular weight of 40,000. Calcium competes with terbium for these binding sites. The affinity for terbium is the same in the intact molecule as in the subunit (KTb ? 19 ± 1 μM), but the affinity for calcium decreases by a factor of 4 (KCa ? 4 mM) in the subunit. Hydrolysis of the receptor, catalyzed by trypsin and chymotrypsin, to peptides with an apparent molecular weight of 8000 or less, does not affect the terbium-binding sites. These experiments indicate that the binding sites for neural activators and for calcium are associated with the same subunit, and that the terbium- and calcium-binding sites reflect structural properties of the polypeptide chain rather than the three-dimensional structure of the protein. 相似文献
9.
Molecular characterization of a stable Flac plasmid 总被引:2,自引:0,他引:2
F L Macrina E Balbinder A Bassel 《Biochemical and biophysical research communications》1973,54(2):737-743
FS is a thermostable extrachromosomal element isolated in which is altered in its replication as compared to its precursor Fts114. Sedimentation of both these plasmids in alkaline sucrose gradients has indicated a difference in their sizes. Contour length measurements of open circular plasmid DNA molecules photographed in the electron microscope have revealed the estimated molecular weight of Fts114 to be 81 × 106 daltons while that of FS is 109 × 106 daltons. FS may carry a segment of chromosomal or cryptic plasmid DNA. 相似文献
10.
Nucleotide sequence of the genes coding for alpha, beta and gamma subunits of the proton-translocating ATPase of Escherichia coli 总被引:12,自引:0,他引:12
H Kanazawa T Kayano K Mabuchi M Futai 《Biochemical and biophysical research communications》1981,103(2):604-612
A nucleotide sequence of 2328 base pairs comprising a portion of the gene cluster for the proton-translocating ATPase of was determined. The sequence covers most of the gene for α subunit, the entire gene for γ subunit and the amino terminal portion of the gene for β subunit, along with the flanking regions of these genes. The amino acid sequences of these subunits deduced from the DNA sequences indicate that the α and γ subunits have 513 and 287 amino acid residues, respectively. A possible secondary structure for each subunit was estimated from the inferred primary structure. The intercistronic regions between the genes for α and γ and between γ and β are 49 and 26 base pairs, respectively. The significance of codon usage in these genes is discussed in correlation with their expression. 相似文献
11.
The complete amino acid sequence of human spleen apoferritin has been determined. It consists of 174 amino acids, corresponding to Mr20017. The sequence is very similar to that of horse spleen apoferritin (14% difference between the two sequences). Some peptides were isolated and sequenced which could not be placed in the sequence but which are homologous with part of the principal sequence. Automatic sequence determination of a large peptide resulting from acid cleavage allows us to establish the presence of two homologous sequences (in the ratio ). 相似文献
12.
The binding of [14C]tuberactinomycin O, an antibiotic closely related to viomycin, to ribosomes has been examined by equilibrium dialysis method. The antibiotic has been observed to bind to the 70S ribosome, which possesses two binding sites: one on the 30S ribosomal subunit and another on the 50S subunit. The affinity for the large subunit is greater than that for the small subunit. The binding to both ribosomal subunits is reversed by viomycin, indicating that tuberactinomycin O and viomycin have the same binding sites on the ribosome. The results seem to be in accordance with the previous finding that viomycin exhibits dual actions on ribosomal function: the inhibition of fMet-tRNAF (initiation) and inhibition of translocation of peptidyl-tRNA. 相似文献
13.
S N Vinogradov S L Hersey R Shukuya 《Biochemical and biophysical research communications》1975,66(2):505-513
The dissociation of the erythrocruorin of the oligochaete was investigated using polyacrylamide gel electrophoresis at neutral pH. In the presence of 0.1% SDS, the erythrocruorin dissociated into five subunits possessing molecular weights of 13,000 (1), 20,000 (2), 23,000 (3), 25,000 (4) and 47,000 (5). In the presence of SDS and mercaptoethanol, the erythrocruorin dissociated into two subunits, whose molecular weights were 13,000 (I) and 28,000 (II). Subunit I accounts for 70–80% of the whole molecule. SDS electrophoresis of the isolated subunits 1 through 5 in the presence of mercaptoethanol showed that subunit I was derived from both subunits 1 and 5, while subunit II was derived from subunits 2–4. These results suggest that erythrocruorin consists of at least five polypeptide chains: two chains of 13,000 and three chains of 28,000. 相似文献
14.
Wen-Tah Hsu 《Biochemical and biophysical research communications》1973,52(3):974-979
The effect of T4 phage on ribosomes in terms of their ability to bind RNA viral template is examined. It is found that the 30S subunits of T4 ribosomes bind MS2 RNA as efficiently as do the subunits of uninfected ribosomes. On the other hand, analyses of the formation of 70S initiation complex, presumably from MS2 RNA-30S ribosome complex, using both labeled MS2 RNA and initiator tRNA, reveal that T4 ribosomes are only about half as active as ribosomes. The latter phenomenon has been reported previously. These results suggest that, following T4 infection, ribosomes are modified in such a way that the attachment of fMet-tRNAf to MS2 RNA-30S subunit complex is impaired. 相似文献
15.
The F1-ATPase from Mycobacterium phlei is inactivated by dicyclohexylcarbodiimide (DCCD), 7-chloro-4-nitrobenzo-2-oxa-1,3-diazole (NBD-Cl) and quinacrine mustard. The inactivation is both time-and concentration-dependent and in the case of DCCD being more pronounced at acidic pH. The minimum inactivation half-time () for DCCD, NBD-Cl and quinacrine mustard was observed to be 14, 6 and 7 min, respectively. Inactivation of F1-ATPase resulted in the incorporation of [14C]DCCD as well as [14C]NBD-Cl into α and γ subunits. The incorporation of label into α and γ subunits, utilizing [14C]NBD-Cl, was reversible by dithiothreitol. Complete inactivation, by linear extrapolation to zero activity, revealed that 4 mol [14C]DCCD and 4 mol [14C]NBD-Cl bind per mol F1-ATPase. Kinetic and binding studies show that these probes bind to site(s) distinct from ATP-binding site in F1-ATPase from M. phlei. 相似文献
16.
Action of methanol on the assocation of ribosomal subunits and its effect on the GTPase activity of elongation factor G 总被引:1,自引:0,他引:1
Methanol causes association of 30S and 50S ribosomal subunits from at MgCl2 concentrations in which they are normally completely dissociated. The 70S ribosome formed under these conditions shows a lower sedimentation velocity and is functionally active in the EF-G GTPase. Association of ribosomal subunits in the presence as well as absence of methanol is affected by washing the ribosomes with 0.5 M NH4Cl. Methanol reduces the Mg2+ concentration required for subunit association as well as for EF-G GTPase activity. The basic requirement for EF-G GTPase activity both with and without alcohol is shown to be the association of 30S and 50S subunits. 相似文献
17.
M R Hanley 《Biochemical and biophysical research communications》1978,82(1):392-401
The phospholipolytic neurotoxin from , crotoxin, is able to produce a dose- and time-dependent block of carbachol-stimulated 22Na efflux from pre-loaded excitable vesicles. The blocking activity is dependent on calcium and is abolished by chemical modification with p-bromophenacyl bromide. The isolated basic subunit, crotoxin B, produces an identical block, whereas the isolated acidic subunit, crotoxin A, has no detectable effect. Neither crotoxin nor crotoxin B antagonizes the binding of to purified acetylcholine receptor, although, at high concentrations, they antagonize its binding to acetylcholine receptor-rich membrane fragments. Certain phospholipase A2 enzymes and the fatty acid products of their digestion can mimic the crotoxin action. It is therefore suggested that, although considered a pre-synaptic neurotoxin, crotoxin can have post-synaptic effects, possibly mediated by its endogeneous phospholipase A2 activity. 相似文献
18.
Hiroaki Tokimatsu William A. strycharz Albert E. Dahlberg 《Journal of molecular biology》1981,152(2):397-412
Three forms of the 50 S ribosomal subunit of Escherichia coli have been separated by agarose/acrylamide gel electrophoresis. The slowest migrating form, S-50 S, corresponded to native 50 S subunits and contained four copies of proteins . Removal of the four copies of this protein produced a more rapidly migrating form, M-50 S. The M-50 S form was then converted to the fastest migrating form, F-50 S, by removal of additional proteins, including L10 and L11. A one-step removal of a pentameric complex of four copies of plus L10 converted the S-50 S subunit directly to the F-50 S subunit. These proteins recombined specifically with the appropriate protein-deficient 50 S subunit at 3 °C to reform the S-50 S subunit, i.e. the M-50 S subunit was converted back to the S-50 S form by the addition of purified proteins ; and the F-50 S subunit bound the pentameric complex of and L10 to form S-50 S. The binding of the pentameric complex, isolated by glycerol gradient centrifugation, supports the model that all four copies of proteins are together in one part of the ribosome called the “ stalk”. Only the four copies of were removed from the 50 S subunit in low salt (0.125 m-NH4Cl) plus 50% ethanol at 0 °C. These ribosomes (in the M-50 S form) had less than 5% of the peptide-synthesizing activity of untreated control ribosomes as measured by a poly(U) translation system in vitro. Peptide-synthesizing activity was restored, upon addition of , back to the treated ribosomes to give 50 S subunits (S-50 S) with a full complement of four copies of . Antibody to proteins bound only to the S-50 S subunits, producing four new bands separated by gel electrophoresis. The bands represented complexes of one, two, three and four antibodies bound to a 50 S subunit. This result was obtained using either 50 S subunits or 70 S tight couples and indicated that all four copies of are either located at a single site in the stalk or, much less likely, are divided between two symmetrical sites. Proteins were not only accessible to their specific antibody but could also be removed from 70 S ribosomes and polyribosomes without causing their dissociation into subunits. The ribosomes and polyribosomes had an increased gel electrophoretic mobility which was reversed by addition of proteins . 相似文献
19.
W.H.M. Peters A.M.M. Fleuren-Jakobs J.J. Schrijen J.J.H.H.M. De Pont S.L. Bonting 《生物化学与生物物理学报:生物膜》1982,690(2):251-260
(1) A preparation from porcine gastric mucosa is solubilized in sodium dodecyl sulfate, and is subjected to gel filtration. (2) A main subunit fraction is obtained, which is a protein carbohydrate lipid complex, containing 88% protein, 7% carbohydrate and 5% phospholipid. The detailed composition of the protein and carbohydrate moieties are reported. (3) Sedimentation analysis of the subunit preparation, after detergent removal, reveals no heterogeneity, but the subunits readily undergo aggregation. (4) Acylation of the subunit preparation with citraconic anhydride causes a clear shift of the band obtained after SDS gel electrophoresis, but the absence of broadening and splitting of the band pleads against subunit heterogeneity. (5) Treatment of the subunit preparation with dansyl chloride indicates that the NH2 terminus is blocked, which favors the assumption of homogeneity of the protein. (6) Binding studies with concanavalin A indicate that at least 86% of the subunit preparation is composed of glycoprotein. (7) These findings, taken together, strongly suggest that there is a single subunit which is a glycoprotein and which represents the catalytic subunit of the enzyme. From sedimentation equilibrium analysis a molecular mass value of 119 kDa (S.E. 3, ) is calculated for protein + carbohydrate and of 110 kDa (S.E. 3, ) for protein only. (8) In combination with the molecular mass of 444 kDa (S.E. 10, ) obtained for the intact enzyme by radiation inactivation we conclude that the enzyme appears to be composed of a homo-tetramer of catalytic subunits. 相似文献
20.
Seung-ki Lee Richard A. Jungmann 《Biochemical and biophysical research communications》1981,102(1):538-544
The phosphorylation of RNA polymerase II after isoproterenol stimulation of confluent rat C6 glioma cell cultures has been investigated. Glioma cells were incubated in the presence of Na2H32PO4 and stimulated for 1 hour with the β-adrenergic agonist isoproterenol. The phosphorylation pattern was analyzed after purification of RNA polymerase II by immunoprecipitation, SDS-polyacrylamide gel electrophoresis and autoradiography. Isoproterenol markedly increased [32P]phosphate incorporation into the 214,000 dalton RNA polymerase subunit. Analysis of the phosphate acceptor amino acid revealed the presence of only [32P]phosphoserine. The data demonstrates an isoproterenol-induced structural modification of RNA polymerase II. 相似文献