Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:Q07644 (polypeptide)
 72,197 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

In a purified system from Escherichia coli containing ribosomes complexed with poly(uridylic acid) and N-acetyl-phenylalanyl-tRNA, the nonhydrolyzable analog of GTP, guanyl-5'-yl imidodiphosphate (Guo-5'-P2-NH-P), promotes polypeptide synthesis at a rate several times slower than GTP. The activity is completely dependent on elongation factors EF-T (i.e, EF-Ts + EF-Tu) and EF-G. Examination of individual steps of the elongation cycle in partial reactions shows that Guo-5'-P2-NH-P is as efficient as GTP in promoting the EF-T-dependent binding of phenylalanyl-tRNA to the ribosomal A site. In contrast, Guo-5'-P2-NH-P promotes the translocation-dependent binding of phenylalanyl-tRNA to a ribosome complexed with A-site-bound N-acetyl-phenylalanyl-tRNA much more slowly than GTP. This slow rate of binding is due to the presence of EF-G on the ribosome, and not to sluggish translocation, since (a) the rate remains slow even after translocation of N-acetylphenylalanyl-tRNA is completed, (b) it is greatly speeded up by removal of EF-G from the reaction mixture (after translocation has occurred), and (c) it is slowed down again by readdition of the factor. Moreover, with post-translocated ribosomes and in the absence of EF-G, formation of dipeptide subsequent to the EF-T-dependent binding of phenylalanyl-tRNA is much slower when binding of this substrate has been promoted by Guo-5'-P2-NH-P than it is when promoted by GTP. The results suggest that, during polymerization with Guo-5'-P2-NH-P, EF-G and EF-Tu are slowly released from the ribosome and, consequently, the steps of the elongation cycle subsequent to translocation and aminoacyl-tRNA binding (aminoacyl-tRNA binding and peptide bond formation, respectively) are delayed. Thus, durong elongation cycle, GTP hydrolysis is probably essential for fast release of the factors from the ribosome.
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PMID:Polypeptide-chain elongation promoted by guanyl-5'-yl imidodiphosphate. 78 22

Escherichia coli phage Qbeta RNA replicase, an RNA-dependent RNA polymerase (RNA-dependent RNA nucleotidyltransferase), is a tetramer composed of one phage-coded polypeptide and three host-supplied polypeptides which are known to function in the biosynthesis of proteins in the uninfected host. Two of these polypeptides, protein synthesis elongation factors EF-Tu and EF-Ts, can be covalently crosslinked with dimethyl suberimidate to form a complex which lacks the ability to catalyze the known host functions catalyzed by the individual elongation factors. Using a previously developed reconstitution system we have examined the effects of crosslinking the EF-Tu-Ts complex on reconstituted replicase activity. Renaturation is significantly more efficient when exogenously added native EF-Tu-Ts is crosslinked than when it is not. Crosslinked EF-Tu-Ts can be purified from a crude crosslinked postribosomal supernatant by its ability to replace EF-Tu and EF-Ts in the renaturation of denatured Qbeta replicase. A sample of Qbeta replicase with crosslinked EF-Tu-Ts replacing the individual elongation factors was prepared. Although it lacked EF-Tu and EF-Ts activities, it could initiate transcription of both poly(C) and Qbeta RNA normally and had approximately the same specific activity as control enzyme. Denatured Qbeta replicase formed with crosslinked EF-Tu-Ts was found to renature much more rapidly than untreated enzyme and, in contrast to normal replicase, its renaturation was not inhibited by GDP. The results demonstrate that EF-Tu and EF-Ts function as complex in Qbeta replicase and do not perform their known protein biosynthetic function in the RNA synthetic reaction.
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PMID:Reconstitution of Qbeta RNA replicase from a covalently bonded elongation factor Tu-Ts complex. 106 92

The interaction of protein synthesis elongation factor 1 (EF-1) from wheat embryos and elongation factor Tu from Escherichia coli with cytidylyl(5'-3')guanosine 5'-triphosphate(pppGpC) has been studied. The dinucleotide 5'-triphosphate interacts strongly with EF-1 as evidenced by its capacity to inhibit the binding of [3H]GTP to the factor. The analogs pGpC and GpC do not interfere with GTP binding to EF-1 but guanosine 5'-triphosphate cyclic 2',3'-monophosphate and ppGpC are also potent inhibitors. The binding of the dinucleotide 5'-triphosphate to EF-1 was also demonstrated directly by the nitrocellulose retention method and by Sephadex G-50 fractionation using a radioactive analog iodinated with 125I in the 5 position of the cytosine of pppGpC. The dinucleotide triphosphate can replace GTP in the formation of a ternary complex EF-1-aminoacyl-tRNA-GTP and in its requirement for the binding of aminoacyl-tRNA to ribosomes catalyzed by EF-1. The absolute requirement for GTP in an in vitro polypeptide-synthesizing system can also be met by pppGpC and by guanosine 5'-triphosphate cyclic 5',3'-monophosphate. The bacterial factor EF-Tu differs drastically from eukaryotic EF-1 in its nucleotide specificity since EF-Tu only interacts slightly (if at all) with pppGpC. The low inhibition of [3H]GTP binding to EF-Tu by pppGpC could be due to a slight contamination in the latter compound.
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PMID:The activity of oligonucleotides containing guanosine 5'-triphosphate in protein synthesis. I. The interaction of protein synthesis elongation factor I with cytidylyl (5'-3')-guanosine 5'-triphosphate. 109 Jun 16

Escherichia coli Phage Qbeta RNA replicase, an RNA-dependent RNA polymerase, is a tetramer composed of one phage-coded polypeptide and three host-supplied polypeptides which are known to function in the biosynthesis of proteins in the uninfected host. Two of these polypeptides, protein synthesis elongation factors EF-Tu and EF-Ts, are required for initiation of transcription by Qbeta replicase with all templates. Using a previously developed reconstitution system we have examined the effects of modification of EF-Tu on reconstituted replicase activity. The poly(G) polymerase activity of the enzyme can be recovered after pretreatment of the EF-Tu-GDP with either L-1-tosylamido-2-phenylethyl chloromethyl ketone or N-ethylmaleimide, both of which inhibit the aminoacyl-tRNA binding activity of EF-Tu. This suggests that the aminoacyl-tRNA binding site of EF-Tu is not required for Qbeta replicase activity. When Qbeta replicase is treated with kirromycin, an antibiotic which modifies EF-Tu activity by an unknown mechamism, the protein synthetic activity of the EF-Tu in the replicase complex is eliminated but the Qbeta RNA replication activity is only slightly affected. Treatment of pure EF-Tu with kirromycin, however, prevents it from functioning in the renaturation of Qbeta replicase. This antibiotic is not effective against the EF-Tu-Ts complex in the reconstitution assay. Kirromycin at the relatively high concentration used here is found to prevent the formation of the EF-Tu-Ts complex. GDP, which binds to EF-Tu and inhibits formation of the complex with EF-Ts, also inhibits renaturation of Qbeta replicase. It is suggested that the EF-Tu-Ts complex, rather than the individual polypeptides, functions in the renaturation of Qbeta replicase and that the kirromycin and GDP act by preventing formation of this complex.
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PMID:Function and structure in ribonucleic acid phage Qbeta ribonucleic acid replicase. Effect of inhibitors of EF-Tu on ribonucleic acid synthesis and renaturation of active enzyme. 126 42

The activity, stability and structure in solution of polypeptide elongation factor hEF-Tu from Halobacterium marismortui have been investigated. The protein is stable in aqueous solutions only at high concentrations of NaCl, KCl or ammonium sulphate, whereas it is more active in exchanging GDP at lower salt concentrations. It is more active and stable at lower pH values than is non-halophilic EF-Tu. The structure in solution of the protein was determined by complementary density, ultracentrifugation, dynamic light-scattering and neutron-scattering measurements. The protein has large hydration interactions, similar to those of other halophilic proteins: 0.4 (+/- 0.1) g of water and 0.20 (+/- 0.05) g of KCl associated with 1 g of protein, with a water/KCl mass ratio always remaining close to 2. The kinetics of inactivation at low salt concentrations showed a stabilizing effect of NaCl when compared to KCl. At low salt concentration, inactivation, protein unfolding and aggregation were strongly correlated. The results suggest that the stabilization model proposed for halophilic malate dehydrogenase by Zaccai et al., involving extensive protein interactions with hydrated salt ions, is also valid for hEF-Tu.
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PMID:Solution studies of elongation factor Tu from the extreme halophile Halobacterium marismortui. 173 Oct 81

Elongation factor EF-1 from Guerin epithelioma was separated into two subunit forms EF-1A and EF-1B by chromatography in the presence of 25% glycerol, successively on CM-Sephadex and DEAE-Sephadex. It was shown that EF-1A is a thermolabile, single polypeptide which catalyses the binding of aminoacyl-tRNA to ribosomes, similarly as eukaryotic EF-1 alpha or prokaryotic EF-Tu. EF-1B was characterized as a complex composed of at least two polypeptides. One of them is EF-1A, the other EF-1C, which stimulates EF-1A activity and protects this elongation factor from thermal inactivation.
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PMID:Purification and properties of the heterogeneous subunits of elongation factor EF-1 from Guerin epithelioma cells. 179 94

The sequence of a 2657 bp DNA fragment containing the coding and regulatory regions of the oxytetracycline (OTC)-resistance gene, otrA, from the OTC producer Streptomyces rimosus was determined. The predicted amino acid sequence of OtrA had extensive identity with tetracycline-resistance genes from other bacteria which mediate resistance via non-covalent ribosomal modification. The N-terminal domain had extremely high identity with the GTP-binding sites of elongation factors, such as EF-G and EF-Tu, suggesting that binding and hydrolysis of GTP is important to the function of the protein. Significant identity with EF-G was present throughout the polypeptide. Transcriptional activity upstream of the otrA coding region was investigated. An Escherichia coli-type promoter, otrAp1, was identified. Transcriptional readthrough of otrA from the upstream gene (otcZ) was also detected in S. rimosus cultures. A divergent promoter activity was identified with subclones of the OtrA fragment in promoter probe vectors analysed in Streptomyces lividans. However, this activity was not identified in a subclone containing more than half of the otrA coding sequence in S. lividans or at all in S. rimosus, indicating that OtrA negatively regulates the expression of the divergent transcript. The data are consistent with regulation of antibiotic production by OtrA to prevent 'suicide'.
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PMID:Characterization of an oxytetracycline-resistance gene, otrA, of Streptomyces rimosus. 180 36

The ras gene product (p21) is a GTP-binding protein and is thought to play an important role in signal transduction of growth and differentiation in many types of mammalian cells. The p21.GTP complex is an active conformation, as described previously for polypeptide chain elongation factors (EF-Tu and EF-G) and heterotrimeric GTP-binding proteins (G proteins). In the study reported here, we measured the amounts of p21-bound guanine nucleotides under various conditions in the G54 cell line, a derivative of Swiss 3T3 cells that overexpresses normal c-Ha-ras. More p21.GTP complexes were present in growing cells than in quiescent cells. When quiescent cells were stimulated with fetal bovine serum to promote DNA synthesis, p21.GTP increased approximately 2-fold. Among a number of purified growth factors, platelet-derived growth factor enhanced the formation of p21.GTP, whereas the combination of bombesin and insulin, which also induces DNA synthesis, did not. These results strongly suggest that p21 is a transducer of the growth signal from the platelet-derived growth factor receptor in Swiss 3T3 cells and that the signal is transmitted through a p21.GTP complex.
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PMID:Platelet-derived growth factor stimulates formation of active p21ras.GTP complex in Swiss mouse 3T3 cells. 219 77

A deletion mutant of a plasmid born Escherichia coli tufA gene, which codes for a truncated elongation factor Tu comprising domains 2 and 3, has been constructed by genetic engineering. This gene was overexpressed in E. coli, and a polypeptide representing the truncated elongation factor Tu was isolated, purified to near homogeneity, crystallized and characterized physico-chemically as well as biochemically. Circular dichroism spectroscopy and limited tryptic digestion demonstrate that the isolated domain pair 2 and 3 behaves like an independent folding unit which adopts a similar secondary and most likely, tertiary, structure to that present in the intact elongation factor Tu. However, the isolated domain pair 2 and 3 does not interact with aminoacyl-tRNA or the antibiotic kirromycin, two ligands which were shown previously by cross-linking experiments to be in contact with amino acid residues located in domains 1 and 2, and domain 3, respectively. The results suggest that the isolated domain pair 2 and 3 by itself forms too few contacts with these ligands to form a stable complex. Furthermore, the data suggest that domain 1 in intact EF-Tu, in a subtle but nevertheless decisive manner, alters the conformation of the other two domains in such a way that all three domains cooperatively create a high affinity binding site for aminoacyl-tRNA and the antibiotic kirromycin.
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PMID:Genetic engineering, isolation and characterization of a truncated Escherichia coli elongation factor Tu comprising domains 2 and 3. 222 77

Transducin, a GTP-binding protein involved in phototransduction in the vertebrate retina, belongs to a family of homologous coupling proteins that also includes Gs and Gi, the regulatory proteins of adenylate cyclase. Here we report the cDNA sequence and deduced amino acid sequence of transducin's alpha subunit (T alpha). The cDNA was isolated, by screening with an antibody probe, from a bovine retinal cDNA library in the expression vector lambda gt11. The 2.2-kilobase cDNA insert hybridized to a single 2.6-kilobase poly(A)+ RNA species present in extracts of bovine retina but not of bovine heart, liver, or brain. The nucleotide sequence of the cDNA revealed an open reading frame long enough to encode the entire 39-kDa T alpha polypeptide. The polypeptide sequence deduced from the cDNA would be composed of 350 amino acids and have a molecular weight of 39,971. Portions of the sequence matched reported amino acid sequences of T alpha tryptic fragments, including sites specifically ADP-ribosylated by cholera and pertussis toxins. The predicted sequence also includes four segments, ranging from 11 to 19 residues in length, that exhibit significant homology to sequences of GTP-binding proteins, including the ras proteins of man and yeast and the elongation factors of ribosomal protein synthesis in bacteria, EF-G and EF-Tu. In combination with previous functional studies of tryptic fragments of T alpha, the deduced amino acid sequence makes it possible to predict which portions of the polypeptide interact with other molecules involved in retinal phototransduction.
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PMID:Amino acid sequence of the alpha subunit of transducin deduced from the cDNA sequence. 240 55


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