Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.7.48 (transcriptase)
 9,479 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

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

In a previous paper we described a number of Escherichia coli mutants resistant to the antibiotic kirromycin. These mutants are altered in both tufA and tufB, the genes coding for elongation factor Tu (EF-Tu). We have now isolated EF-Tu in a homogeneous form from the mutant strains and have studied its function in polypeptide synthesis. These EF-Tu preparations were examined in renaturation studies of Qbeta RNA replicase, described in another paper. In order to characterize the factor we have inactivated the tufB gene by insertion of bacteriophage Mu or by an amber mutation. This enabled us to isolate EF-Tu as a single gene product derived from tufA (designated EF-TuA in contrast to the tufB product, which is called EF-TuB). Kirromycin-resistant EF-TuA did not respond to addition of the antibiotic in three assays: [(3)H]GDP exchange with EF-Tu-GDP at 0 degrees C, in vitro translation of poly(U), and kirromycin-induced GTPase activity of EF-Tu. In contrast, wild-type EF-TuA responded normally to the antibiotic in these assays. One of our mutants (LBE 2012) harbors the kirromycin-resistant EF-TuA and an EF-TuB that is able to bind kirromycin. This binding does not cause inhibition of protein synthesis, indicating that EF-TuB from LBE 2012 is unable to reach the ribosome under these conditions. The two types of EF-Tu from this mutant are equal in size but differ by 0.1 pH unit in isoelectric point. In the soluble fractions of LBE 2012 cells they are present in approximately equal amounts. Our results also show that the tufB gene is not necessary for bacterial growth.
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PMID:Elongation factor Tu isolated from Escherichia coli mutants altered in TufA and tufB. 700 48