共查询到20条相似文献,搜索用时 31 毫秒
1.
Polyoma-infected 3T6 cells contain a number of proteins precipitable by serum from rats carrying polyoma-induced tumors. The virus codes for three species having apparent molecular weights of 90,000, 60,000 and 22,000 daltons, as determined by polyacrylamide gel electrophoresis (90K, 60K and 22k). The 90K and 22K species produced by a large plaque and a small plaque wild-type polyoma have similar mobilities, but the 60K species produced by the large plaque wild-type. In cells infected by each of seven polyoma tsA mutants, the 90K species is unstable at the nonpermissive temperature, while the 60K and 22K species are stable. In cells infected by a mutant carrying a deletion between roughly 98 and 3 map units in the early region of the viral genome, the 22K species is present, but the 90K and 60K species are absent. Tryptic peptide analysis of the isolated 90K, 60K and 22K species shows that the three species have common N terminal regions. The 60K and 22K species contain amino acid sequences not found in the 90K species , and the 60K species has several unique, methionine-containing peptides not found in either the 22K or 90K species. Two polyoma-transformed BHK cell lines do not have detectable amounts of the 90K protein. 相似文献
2.
Structural Proteins of Intracisternal A-Particles: Possible Repetitive Sequences 总被引:3,自引:3,他引:0
下载免费PDF全文
![点击此处可从《Journal of virology》网站下载免费的PDF全文](/ch/ext_images/free.gif)
Peptide maps of tryptic digests of the structural proteins from inner shells of intracisternal A-particles have shown common peptides for all the proteins. The terminal amino group of the three different structural proteins was identified as arginine. The major protein revealed approximately half the number of peptides expected from the amino acid composition. Since evidence for a cross-link bond has not been found, the main structural protein may be a single polypeptide chain containing a total or partial duplication of sequence. 相似文献
3.
4.
Immunological and Chemical Identification of Intracellular Forms of Adenovirus Type 2 Terminal Protein 总被引:3,自引:2,他引:1
下载免费PDF全文
![点击此处可从《Journal of virology》网站下载免费的PDF全文](/ch/ext_images/free.gif)
Maurice Green Janey Symington Karl H. Brackmann Maria A. Cartas Helen Thornton Leann Young 《Journal of virology》1981,40(2):541-550
Highly purified adenovirus type 2 terminal protein (TP) with an apparent Mr of 55,000 (55K) was prepared in quantities of 10 to 30 μg from guanidine hydrochloride- or sodium dodecyl sulfate-disrupted virions (60 to 120 mg). Guinea pigs were immunized with 14 to 20 injections of TP in amounts of 1 to 2 μg. Antiserum to TP was used to study the intracellular polypeptides related to adenovirus type 2 TP. By immunoprecipitation with anti-TP serum, we identified 80K and 76K polypeptides in the nucleoplasmic and cytoplasmic S100 fractions of [35S]methionine-labeled cells early and late after infection with Ad2. By immunoautoradiographic analysis which eliminates coprecipitation of unrelated proteins, we identified an 80K polypeptide (probably an 80K-76K doublet) in unlabeled, late infected cells, using anti-TP serum and 125I-labeled staphylococcal protein A. About two- to threefold-higher levels of the 80K and 76K polypeptides were present in the nucleoplasm than in the S100 fraction, and two- to threefold-higher levels were found in late infected cells than in early infected cells (cycloheximide enhanced, arabinofuranosylcytosine treated). We did not detect the 80K or 76K polypeptide in uninfected cells, indicating that these polypeptides are virus coded. Tryptic peptide map analysis showed that the 80K and 76K polypeptides are very closely related and that they share peptides with the DNA-bound 55K TP. Our data provide the first direct demonstration of intracellular 80K and 76K forms of TP. The intracellular 80K and 76K polypeptides are closely related or identical to the 80K polypeptide that Challberg and co-workers (Proc. Natl. Acad. Sci. U.S.A. 77:5105-5109, 1980) detected at the termini of adenovirus DNA synthesized in vitro and to the 87K polypeptide that Stillman and co-workers (Cell 23:497-508, 1981) translated in vitro. We did not detect the 55K TP in early or late infected cells, consistent with the proposal by Challberg and co-workers that the 80K polypeptide is a precursor to the virion-bound TP and that the conversion of the 80K polypeptide to the 55K TP occurs during virus maturation. The 80K and 76K polypeptides have many more methionine-containing tryptic peptides than does the 55K TP, and most of the tryptic peptides unique to the 80K and 76K polypeptides are very hydrophobic. Thus, the conversion of the 80K and 76K polypeptides to the 55K TP may involve the removal of a specific hydrophobic protein region. 相似文献
5.
The amino acid sequence of ribosomal protein S18 from Bacillus stearothermophilus has been completely determined by automated sequence analysis of the intact protein as well as of peptides derived from digestion with Staphylococcus aureus protease at pH 4.0 and cleavage with cyanogen bromide. The carboxy-terminal region was verified by both amino acid analyses of chymotryptic peptides and by mass spectrometry from the terminal region. The protein contains 77 amino acid residues and has an Mr of 8838. Comparison of this sequence with the sequences of the S18 proteins from tobacco and liverwort chloroplasts and E. coli shows a relatively high similarity, ranging from 42 to 55% identical residues with the B. stearothermophilus S18 protein. The regions of homology common to all four proteins consist of several positively charged sections spanning the entire length of the protein. 相似文献
6.
Identification and characterization of the protein encoded by the human c-myb proto-oncogene. 总被引:18,自引:4,他引:14
下载免费PDF全文
![点击此处可从《The EMBO journal》网站下载免费的PDF全文](/ch/ext_images/free.gif)
We have identified the product of the human c-myb proto-oncogene as a 80,000-Mr protein, p80c-myb, by using polyclonal and monoclonal antibodies raised against a bacterially synthesized polypeptide from the amino terminus of the viral myb protein. p80c-myb shares at least two distinct antigenic sites with the amino terminal region of the v-myb protein. p80c-myb is found only in hematopoietic cells or in cells that contain amplified c-myb genes. Like the chicken myb proteins, p80c-myb is a nuclear DNA-binding protein that is predominantly associated with chromatin and exhibits a short half-life of approximately 1 hour. 相似文献
7.
Four mutants of polyoma virus lacking endonuclease HindII site 1 were isolated and characterized with respect to the VP1 coding sequence. Three of these mutants had deletions that removed 0.2 to 0.3% of the genome. All three deletion mutants encoded VP1 proteins that were smaller than wild type and that lacked one or more tryptic peptides normally found in the wild-type VP1 protein. Our results suggest the HindII site 1 is at, or very near, the carboxy terminal end of the coding sequence for VP1. A model for the peptide organization in that region is presented. 相似文献
8.
Regina Monaco James M. Chen Denise Chung Paul Brandt-Rauf Matthew R. Pincus 《The protein journal》1995,14(6):457-466
Theras-oncogene-encoded p21 protein becomes oncogenic if amino acid substitutions occur at critical positions in the polypeptide chain. The most commonly found oncogenic forms contain Val in place of Gly 12 or Leu in place of Gln 61. To determine the effects of these substitutions on the three-dimensional structure of the whole p21 protein, we have performed molecular dynamics calculations on each of these three proteins bound to GDP and magnesium ion to compute the average structures of each of the three forms. Comparisons of the computed average structures shows that both oncogenic forms with Val 12 and Leu 61 differ substantially in structure from that of the wild type (containing Gly 12 and Gln 61) in discrete regions: residues 10–16, 32–47, 55–74, 85–89, 100–110, and 119–134. All of these regions occur in exposed loops, and several of them have already been found to be involved in the cellular functioning of the p21 protein. These regions have also previously been identified as the most flexible domains of the wild-type protein and have been bound to be the same ones that differ in conformation between transforming and nontransforming p21 mutant proteins neither of which binds nucleotide. The two oncogenic forms have similar conformations in their carboxyl-terminal domains, but differ in conformation at residues 32–47 and 55–74. The former region is known to be involved in the interaction with at least three downstream effector target proteins. Thus, differences in structure between the two oncogenic proteins may reflect different relative affinities of each oncogenic protein for each of these effector targets. The latter region, 55–74, is known to be a highly mobile segment of the protein. The results strongly suggest that critical oncogenic amino acid substitutions in the p21 protein cause changes in the structures of vital domains of this protein. 相似文献
9.
Nucleotide sequence of the gene for the ferrienterochelin receptor FepA in Escherichia coli. Homology among outer membrane receptors that interact with TonB 总被引:48,自引:0,他引:48
We have determined the nucleotide sequence of the Escherichia coli fepA gene, which codes for the outer membrane receptor for ferrienterochelin and colicins B and D. The predicted FepA polypeptide has a molecular weight of 79,908 and consists of 723 amino acids. A 22-amino acid leader or signal peptide preceded the mature protein. With respect to overall composition, hydropathy, net charge and distribution of nonpolar segments, the FepA polypeptide was typical of other E. coli outer membrane proteins, except that FepA contained 2 cysteine residues. Comparison of the deduced amino acid sequence of FepA with that of three other TonB-dependent receptors (BtuB, FhuA, and IutA) revealed only a few regions of sequence homology; one of these included the amino-termini. An amino acid substitution within the conserved amino-terminal region of BtuB resulted in production of a receptor that had normal binding functions but was incapable of energy-dependent transport of vitamin B12. This result suggests that the amino-terminal end of these four polypeptides is involved in interaction with the TonB protein or another step of energy transduction. Three other regions of homology were shared among the four proteins: one near residues 50 to 70, another at about residue 100 to 140, and the last between 20 and 40 amino acid residues from the carboxyl terminus. The function of these three regions remains speculative. 相似文献
10.
Cardiac Na+/Ca(2+)-exchange is an integral membrane protein consisting of approx. 970 amino acids with as many as 12 membrane-spanning and 11 extramembranal regions (Nicoll, D.A., Lognoni, S. and Philipson, K.D. (1985) Science 250, 562-565). Based upon primary sequence information, 3 amino-acid sequences located in either extramembranal segment a or f, consisting largely of acidic amino-acids, were selected for the production of synthetic peptides. The peptides were cross-linked to carrier ovalbumin and used to generate site-directed polyclonal antibodies (sd-Ab). Western blot analysis of bovine cardiac sarcolemmal (SL) proteins demonstrated that sd-Ab against segment a and 1 against loop f recognized a 70 kDa protein and a lower molecular mass band at 55 kDa under reducing conditions. A different loop f sd-Ab failed to recognize the 70 kDa protein but did associate with a 120, 65 and 55 kDa protein under the same conditions. Under non-reducing conditions, antibodies to all three peptides recognized the 65 kDa protein. All sd-Ab were blocked by addition of their respective peptides and were not inhibited by either of the other peptides. A sd-Ab against loop f was immobilized to an affinity support matrix and used to immunoprecipitate detergent solubilized cardiac SL vesicle protein. Immunoprecipitated protein was reconstituted into proteoliposomes which demonstrated Na+/Ca(2+)-exchange activity. Immunoprecipitated protein cross-reacted with sd-Ab against all three peptides with bands at 120, 70 and 55 kDa on Western blots. Tryptic digests of native SL vesicles abolished recognition of segment a sd-Ab for SL proteins while having little or no affect on reactivity to the protein by both sd-Ab against loop f. Digestion of the SL vesicle protein with endoproteinase Arg C did not alter sd-Ab recognition. The results suggest that specific domains of the cardiac Na+/Ca(2+)-exchanger depending upon the conformation of the protein, may not be available for antibody binding. The 70 kDa polypeptide appears to include the N-terminal region of the protein and what is believed to be a large cytoplasmic extramembranal loop. Limited proteolysis by trypsin and endoproteinase Arg C yielded results consistent with the model which places the N-terminus of the protein on the extracellular surface and a large extramembranal segment (loop f) on the cytoplasmic side of the SL membrane. 相似文献
11.
Comparison of protein sulfation in control and virus-transformed baby hamster kidney cells 总被引:2,自引:0,他引:2
Protein sulfation in baby hamster kidney cells (BHK) and their polyoma virus transformants (PY-BHK) was studied comparatively. On in vivo labeling, [35S]-sulfate was incorporated into the 50K protein and proteins in the 100-180K range, represented by the 155K protein. The incorporation into both the 50K and 155K protein was elevated 2-3 fold in PY-BHK cells compared to in BHK cells. Tyrosine-O-sulfate was the only identifiable sulfated amino acid in both proteins. On in vitro labeling with [35S]3'-phosphoadenosine 5'-phosphosulfate (PAPS), at least 6 radioactive protein bands were discernible on gel electrophoresis. Of these, sulfation of the 57K and 60K proteins was elevated in PY-BHK cells compared to in BHK cells, whereas sulfation of the 39K protein was depressed in PY-BHK cells. Tyrosine-O-sulfate was the only identifiable sulfated amino acid in these proteins. 相似文献
12.
Regina Monaco James M. Chen Denise Chung Paul Brandt-Rauf Matthew R. Pincus 《Journal of Protein Chemistry》1995,14(6):457-466
Theras-oncogene-encoded p21 protein becomes oncogenic if amino acid substitutions occur at critical positions in the polypeptide chain. The most commonly found oncogenic forms contain Val in place of Gly 12 or Leu in place of Gln 61. To determine the effects of these substitutions on the three-dimensional structure of the whole p21 protein, we have performed molecular dynamics calculations on each of these three proteins bound to GDP and magnesium ion to compute the average structures of each of the three forms. Comparisons of the computed average structures shows that both oncogenic forms with Val 12 and Leu 61 differ substantially in structure from that of the wild type (containing Gly 12 and Gln 61) in discrete regions: residues 10–16, 32–47, 55–74, 85–89, 100–110, and 119–134. All of these regions occur in exposed loops, and several of them have already been found to be involved in the cellular functioning of the p21 protein. These regions have also previously been identified as the most flexible domains of the wild-type protein and have been bound to be the same ones that differ in conformation between transforming and nontransforming p21 mutant proteins neither of which binds nucleotide. The two oncogenic forms have similar conformations in their carboxyl-terminal domains, but differ in conformation at residues 32–47 and 55–74. The former region is known to be involved in the interaction with at least three downstream effector target proteins. Thus, differences in structure between the two oncogenic proteins may reflect different relative affinities of each oncogenic protein for each of these effector targets. The latter region, 55–74, is known to be a highly mobile segment of the protein. The results strongly suggest that critical oncogenic amino acid substitutions in the p21 protein cause changes in the structures of vital domains of this protein. 相似文献
13.
Organization of early region 1B of human adenovirus type 2: identification of four differentially spliced mRNAs. 总被引:16,自引:10,他引:6
下载免费PDF全文
![点击此处可从《Journal of virology》网站下载免费的PDF全文](/ch/ext_images/free.gif)
The mRNAs from early region 1B of adenovirus type 2 have been studied by Northern blot, S1 nuclease, and cDNA analysis. Two novel mRNAs, designated 14S and 14.5S, have been observed in addition to the previously identified 9S, 13S, and 22S mRNAs. They are 1.26 and 1.31 kilobases long and differ from the 13S and 22S mRNAs in being composed of three exons instead of two. Their two terminal exons are the same as those present in the 13S mRNA, whereas the middle exon is unique to each of the two novel mRNA species. The structures of the 14S and 14.5S mRNAs allow the prediction of their coding capacities: both mRNA species, like the 22S and 13S mRNAs, contain an uninterrupted translational reading frame encoding a 21,000-molecular-weight (21K) polypeptide. The 14S mRNA can, in addition, encode a 16.5K polypeptide which shares N-terminal and C-terminal sequences with the 55K polypeptide, known to be encoded by the 22S mRNA. The 14.5S mRNA species encodes a hypothetical 9.2K polypeptide which has the same N terminus as the 55K polypeptide but a unique C terminus. The two mRNAs differ in their kinetics of appearance; the 14.5S mRNA is preferentially expressed late after infection in contrast to the 14S mRNA, which is present in approximately equal amounts early and late after infection. Taken together with previously published information the results suggest that early region 1B of adenovirus type 2 encodes five proteins in addition to virion polypeptide IX. These have predicted molecular weights of 55,000, 21,000, 16,500, 9,200, and 8,100. 相似文献
14.
15.
Characterization of Rous sarcoma virus src gene products synthesized in vitro. 总被引:98,自引:33,他引:65
下载免费PDF全文
![点击此处可从《Journal of virology》网站下载免费的PDF全文](/ch/ext_images/free.gif)
The cell-free synthesis of three major proteins from virion RNA of nondefective Rous sarcoma virus (RSV), but not from RNA of transformation-defective deletion mutants, has been observed. The apparent molecular weights of these transformation-specific proteins are approximately 60,000 (60K), 25K, and 17K. Tryptic maps of methionine-containing peptides revealed the 17K, 25K, and 60K proteins to be overlapping in sequence. However, only partial homology was observed between the 17K, 25K and 60K proteins synthesized from Schmidt-Ruppin strain, subgroup D, RSV RNA and those synthesized from Prague strain, subgroup B, RSV, RNA. About half of the methionine peptides in the Schmidt-Ruppin strain, subgroup D, 60K protein were shared with the Prague strain, subgroup D, 60K protein, and the rest were distinct to each. The virion RNAs coding for the 60K, 25K, and 17K proteins were found to be polyadenylated and to sediment with maximal mRNA activity at about 23, 19 to 20, and 18S, respectively. In addition, transformation-specific proteins with molecular weights of 39K and 33K were observed by in vitro synthesis. These proteins are also related to the 60K, 25K, and 17K proteins and were synthesized from polyadenylated RSV RNA of approximately 21 to 22S. RNase T1-resistant oligonucleotides were analyzed in parallel, and the src-specific oligonucleotides were found to be first present in equimolar amounts in those gradient fractions sedimenting at 21 to 22S. Our data suggest that synthesis of the 60K protein is initiated near the 5' terminus of the src gene, whereas the 39K, 33K, 25K, and 17K proteins are initiated internally in the src gene. All of these proteins appear to be initiated independently, but they may have a common termination site. 相似文献
16.
Analysis of adenovirus early region 4-encoded polypeptides synthesized in productively infected cells. 总被引:34,自引:25,他引:9
下载免费PDF全文
![点击此处可从《Journal of virology》网站下载免费的PDF全文](/ch/ext_images/free.gif)
Peptide-specific antisera were developed to analyze the products encoded by adenovirus type 5 early region 4 (E4) open reading frames 6 and 7. Reading frame 6 previously was shown to encode a 34-kilodalton polypeptide (34K polypeptide) that forms a complex with the early region 1B (E1B)-55K antigen and is required for efficient viral growth in lytic infection. Antisera that were generated recognized the E4-34K protein as well as a family of related polypeptides generated by the fusion of open reading frames 6 and 7. These polypeptides shared amino-terminal sequences with the 34K protein. Short-pulse analysis suggested that the heterogeneity observed with the 6/7 fusion products resulted from differential splicing patterns of related E4 mRNAs. An antiserum directed against the amino terminus of reading frame 6 recognized only the free form of the 34K antigen that was not associated with the E1B-55K protein. This observation allowed the determination of the stability of the free and complexed form of this polypeptide. Pulse-chase analyses demonstrated that both forms of the 34K protein had half-lives greater than 24 h, suggesting that complex formation did not result in stabilization of this gene product. The half-lives of the 6/7 fusion products were approximately 4 h. The 34K protein also was shown to have a nuclear localization within infected cells. Finally, analysis of a mutant carrying deletions in both the E4-34K and E1B-55K polypeptides indicated that the complex formed between these two proteins was a functional unit in lytic infection. 相似文献
17.
Identification of an autoinhibitory domain in the C-terminal region of the plant plasma membrane H(+)-ATPase. 总被引:12,自引:0,他引:12
M G Palmgren M Sommarin R Serrano C Larsson 《The Journal of biological chemistry》1991,266(30):20470-20475
Proteolytic (trypsin) treatment removes a small terminal segment from the 100-kDa plant plasma membrane H(+)-ATPase. This results in activation of H+ pumping across the plasma membrane, suggesting that an inhibitory domain is located in one of the terminal regions of the enzyme (Palmgren, M.G., Larsson, C., and Sommarin, M. (1990) J. Biol. Chem. 265, 13423-13426). In order to identify the origin of the fragment released by trypsin, polyclonal antibodies were raised against the first 55 amino acids (N-terminal region), the last 99 amino acids (C-terminal region), and a portion of 150 amino acids in the central part of the enzyme as deduced from one of the H(+)-ATPase genes (PMA2) of Arabidopsis thaliana. The native, 100-kDa H(+)-ATPase was recognized by all three antisera in Western blots. By contrast, the approximately 90-kDa polypeptide appearing after trypsin treatment was only recognized by the antisera against the N-terminal and central region, but not by the antiserum against the C-terminal region, suggesting that the inhibitory domain is located in this part of the enzyme. To more closely determine the position of the inhibitory domain, three peptides representing conserved parts of the C-terminal region were synthesized (residues 861-888, 912-943, and 936-949 of the Arabidopsis (PMA2) sequence). Only one of the peptides (residues 861-888) affected H+ pumping by the trypsin-activated (approximately 90-kDa) enzyme. This peptide of 28 amino acids inhibited H+ pumping with an IC50 of about 15 microM, suggesting that the auto-inhibitory domain is located within the corresponding part of the C-terminal region. 相似文献
18.
D Barbeau R C Marcellus S Bacchetti S T Bayley P E Branton 《Biochimie et biologie cellulaire》1992,70(10-11):1123-1134
Human adenovirus E1A proteins and oncogene products of several other DNA tumour viruses derive much of their oncogenic potential from interactions with cellular polypeptides. E1A proteins form complexes with p105Rb and a related p107 polypeptide, and with at least three other proteins (p60cycA, p130, and p300); all may be required for cell transformation. Using a series of E1A deletion mutants, we have carried out a quantitative analysis of the binding patterns of cellular proteins to E1A products. Binding of most of the proteins was affected at least partially by mutations within the amino terminal 25 residues, amino acids 36-69 within conserved region 1 (CR1), and residues 121-138 in conserved region 2 (CR2). However, the specific binding characteristics of each protein varied considerably. p300 was the only species for which binding was totally eliminated by deletions at the amino terminus. Removal of regions within CR1 eliminated binding of all species except p107 and p60cycA. Deletion of portions of CR2 reduced or eliminated binding of all proteins except p300. Thus, whereas cellular polypeptides generally were found to interact with the same three regions of E1A proteins, specific interactions varied considerably. 相似文献
19.
N C Franklin 《Journal of molecular biology》1985,181(1):85-91
20.
Identification of sites influencing the folding and subunit assembly of the P22 tailspike polypeptide chain using nonsense mutations 总被引:4,自引:2,他引:2
下载免费PDF全文
![点击此处可从《Genetics》网站下载免费的PDF全文](/ch/ext_images/free.gif)
Amber mutations have been isolated and mapped to more than 60 sites in gene 9 of P22 encoding the thermostable phage tailspike protein. Gene 9 is the locus of over 30 sites of temperature sensitive folding (tsf) mutations, which affect intermediates in the chain folding and subunit association pathway. The phenotypes of the amber missense proteins produced on tRNA suppressor hosts inserting serine, glutamine, tryosine and leucine have been determined at different temperatures. Thirty-three of the sites are tolerant, producing functional proteins with any of the four amino acids inserted at the sites, independent of temperature. Tolerant sites are concentrated at the N-terminal end of the protein indicating that this region is not critical for conformation or function. Sixteen of the sites yield temperature sensitive missense proteins on at least one nonsense suppressing host. Most of the sites with ts phenotypes map to the central region of the gene which is also the region where most of the tsf mutations map. Mutations at 15 of the sites have a lethal phenotype on at least one tRNA suppressor host. For nine out of ten sites tested with at least one lethal phenotype, the primary defect was in the folding or subunit association of the missense polypeptide chain. This analysis of the tailspike missense proteins distinguishes three classes of amino acid sites in the polypeptide chain; residues whose side chains contribute little to folding, subunit assembly or function; residues critical for maintaining the folding and subunit assembly pathway at the high end of the temperature range of phage growth; and residues critical over the entire temperature range of growth. 相似文献