Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

---

Query: EC:2.7.7.6 (RNA polymerase)
34,946 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The initiator AUC of the mouse dihydrofolate reductase gene (dhfr) was converted to ACG by site-directed mutagenesis and assayed for expression in cultured monkey cells using an SV40 recombinant called SVGT5dhfr26m2. Synthesis of apparently full-length dihydrofolate reductase (DHFR) protein was significantly reduced compared to wild-type, but not entirely abolished, suggesting that the ACG triplet was being utilized for translation initiation. In addition, a truncated form of DHFR was produced, apparently by initiation at the next in-frame AUG downstream. This result was confirmed in vitro. Transcripts of the dhfr sequence were produced by SP6 RNA polymerase in the presence of m7GpppG and translated in vitro using reticulocyte lysates and wheat germ extracts. The results paralleled those observed in vivo. Synthesis of full-length DHFR was reduced, but not eliminated, and a new species was produced by initiation at an internal site. Amino acid sequence analysis of the products of in vitro translation demonstrated that translation does indeed initiate at the ACG triplet and that it initiates with methionine. Additional mutations were introduced which altered the sequence context of the ACG triplet. Mutation of the translation initiation consensus sequence by substitution of the A residue at position -3, or of the G at +4 resulted in a significant decrease in initiation at the ACG and an increase in the level of the internal initiation product. Thus, translation initiation at a non-AUG triplet depends on a favorable sequence context.
...
PMID:Translation initiation at an ACG triplet in mammalian cells. 304 Jul 20

The gene expression of nine phages of the T7 group was compared after infection of Escherichia coli B(P1). With the exception of phage 13a which grew normally, all of them infected E. coli B(P1) abortively. Differences were found in the efficiency of host killing which ranged from 100% for phage 13a to 37% for phage A1122. Infection by T7 prevented colony formation by about 70% of the cells but they showed filamentous growth until about 2 h after infection. It was shown by SDS-polyacrylamide gel electrophoresis and autoradiography of [35S]methionine-labelled phage-coded proteins that all phages except for 13a showed measurable expression only of the early genes. No correlation was observed between killing capacity and the pattern of gene expression, and the ability to hydrolyse S-adenosyl-methionine (SAM, a cofactor for the P1 restriction endonuclease) by means of a phage-coded SAMase. Mixed infection of E. coli B(P1) with 13a and T7 yielded mixed progeny indistinguishable from that observed after mixed infection of the normal host E. coli B. Genetic crosses with amber mutants of 13a and T7 showed that the 13a marker opo+ (overcomes P one), required for growth on B(P1), is located in the early region, to the left of gene 1 (RNA polymerase gene).
...
PMID:Inhibition of gene expression of T7-related phages by prophage P1. 304 52

Citrate synthase is a key enzyme of the Krebs tricarboxylic acid cycle and catalyzes the stereospecific synthesis of citrate from acetyl coenzyme A and oxalacetate. The amino acid sequence and three-dimensional structure of pig citrate synthase dimers are known, and regions of the enzyme involved in substrate binding and catalysis have been identified. A cloned complementary DNA sequence encoding pig citrate synthase has been isolated from a pig kidney lambda gt11 cDNA library after screening with a synthetic oligonucleotide probe. The complete nucleotide sequence of the 1.5-kilobase cDNA was determined. The coding region consists of 1395 base pairs and confirms the amino acid sequence of purified pig citrate synthase. The derived amino acid sequence of pig citrate synthase predicts the presence of a 27 amino acid N-terminal leader peptide whose sequence is consistent with the sequences of other mitochondrial signal peptides. A conserved amino acid sequence in the mitochondrial leader peptides of pig citrate synthase and yeast mitochondrial citrate synthase was identified. To express the pig citrate synthase cDNA in Escherichia coli, we employed the inducible T7 RNA polymerase/promoter double plasmid expression vectors pGP1-2 and pT7-7 [Tabor, S., & Richardson, C. C. (1985) Proc. Natl. Acad. Sci. U.S.A. 82, 1074-1078]. The pig citrate synthase cDNA was modified to delete the N-terminal leader sequence; then by use of a synthetic oligonucleotide linker, the modified cDNA was cloned into pT7-7 immediately following the initiator Met. A glutamate-requiring (citrate synthase deficient), recA- E. coli mutant, DEK15, was transformed with pGP1-2 and then pT7-7PCS. pT7-7PCS complemented the E. coli gltA mutation.(ABSTRACT TRUNCATED AT 250 WORDS)
...
PMID:Isolation, nucleotide sequence, and expression of a cDNA encoding pig citrate synthase. 304 87

The regulation of ornithine decarboxylase (ODC) activity by the polyamine derivatives N1,N8-bis(ethyl)-spermidine and N1,N12-bis(ethyl)spermine was studied using a line of L1210 cells resistant to alpha-difluoromethylornithine (D-R cells), which contain very high levels of ODC, and a synthetic mRNA prepared from a plasmid containing an insert corresponding to ODC mRNA adjacent to an SP6 RNA polymerase promoter. Studies in which ODC protein was labeled in the D-R cells by exposure to [35S]methionine indicated that the polyamine derivatives and their physiological counterparts led to an increased rate of degradation of ODC and to a rapid reduction in ODC synthesis without affecting the content of ODC mRNA. Direct evidence that the polyamine derivatives act by inhibiting the translation of the ODC mRNA was obtained by studying their effects on the translation of ODC mRNA in reticulocyte lysates. This translation was strongly inhibited by the addition of N1,N8-bis(ethyl)spermidine, spermidine, N1,N12-bis(ethyl)spermine, or spermine but was not affected much by putrescine. The inhibition of the translation of ODC mRNA by either of the bis(ethyl) polyamine derivatives occurred at concentrations which stimulated total protein synthesis showing the selectivity of the reduction in ODC. The effects of polyamine derivatives and polyamines on translation of the plasmid-derived ODC mRNA were identical with those found with the D-R L1210 cell mRNA. This synthetic ODC mRNA lacks 261 bases of the 5'-leader sequences and 200 bases plus the poly(A) section from the 3'-nontranslated sequence. Therefore, these regions appear not to influence sensitivity of the ODC mRNA to inhibition of translation by polyamine derivatives.
...
PMID:Control of ornithine decarboxylase activity in alpha-difluoromethylornithine-resistant L1210 cells by polyamines and synthetic analogues. 313 56

A series of specific deletion mutants derived from a full-length cDNA clone of cowpea mosaic virus (CPMV) B RNA was constructed with the aim to study the role of viral proteins in the proteolytic processing of the primary translation products. For the same purpose cDNA clones were constructed having sequences derived from both M and B RNA of CPMV. In vitro transcripts prepared from these clones with T7 RNA polymerase, were efficiently translated in rabbit reticulocyte lysates. The translation products obtained were processed in the lysate by specific proteolytic cleavages into smaller products, which made it possible to study subsequently the effect of the various mutations on this process. The results obtained indicate that the B RNA-encoded 24K polypeptide represents a protease responsible for all cleavages in the polyproteins produced by both CPMV B and M RNA. For efficient cleavage of the glutamine-methionine site in the M RNA encoded polyprotein the presence of a second B RNA encoded protein, the 32K polypeptide, is essential, although the 32K polypeptide itself does not have proteolytic activity. A number of cleavage-site mutants were constructed in which the coding sequence for the glutamine-glycine cleavage site between the two capsid proteins was changed. Subsequent in vitro transcription and translation of these cleavage site mutants show that a correct dipeptide sequence is a prerequisite for efficient cleavage but that the folding of the polypeptide chain also plays an important role in the formation of a cleavage site.
...
PMID:Two viral proteins involved in the proteolytic processing of the cowpea mosaic virus polyproteins. 328 25

An in vitro mixed transcription system was employed to examine the possible alteration of the promoter selectivity of Escherichia coli RNA polymerase by specific tRNAs. Transcription in vitro was inhibited by most of the tRNAs examined, although the extent of the inhibition differed with the tRNA species. The inhibition by tRNAs was due to competition with DNA for binding RNA polymerase. This inhibitory effect remained after charging of the tRNAs with amino acids. The charging of tRNAfMet with fMet, but not with Met, abolished its inhibitory effect, and instead gave a stimulatory effect on the transcription from some promoters. These observations suggest that fMet-tRNAfMet plays a specific regulatory role in the coupling of transcription to translation.
...
PMID:Promoter selectivity of Escherichia coli RNA polymerase: alteration by fMet-tRNAfMet. 353 31

Transcription in vitro by the RNA polymerase of infectious haematopoietic necrosis virus (IHNV), a salmonid rhabdovirus, was investigated using different reaction conditions to maximize RNA synthesis. The use of HEPES buffer rather than Tris buffer, and the addition of S-adenosyl-L-methionine to the reactions resulted in a sixfold increase in RNA synthetic activity to 6400 pmol UMP incorporated/mg viral protein/hour. The RNA transcripts produced in this system contained polyadenylated species which co-migrated with IHNV mRNA species 2, 3, 4 and 5 from IHNV-infected cells. The transcripts were shown to be functional mRNA species by their ability to direct the synthesis of viral proteins in vitro.
...
PMID:Transcription in vitro of infectious haematopoietic necrosis virus, a fish rhabdovirus. 358 84

[35S]Methionine-labeled proteins from total or cytoplasmic extracts of Vero cells infected with African swine fever virus were chromatographed on native and denatured DNA-cellulose and DNA-binding proteins were analyzed by SDS-polyacrylamide gel electrophoresis (SDS-PAGE), by DNA binding to Western blots, or by two-dimensional electrophoresis. Thirteen virus-specific DNA-binding proteins were detected by one-dimensional analysis. Major species have molecular mass 44,000 (44K), 38K, 20K, 18K, 14K, 13K, and 12K. The remaining DNA-binding proteins are proteins with molecular mass 130K, 110K, 35K, 33K, 17K, and 14.5K. When viral DNA used in the binding assay the results were very similar but the 13K protein did not bind viral DNA. Seven other minor virus-specific DNA-binding proteins could be detected by two-dimensional analysis. This technique also enabled the assignment of virus-specific proteins. Seven of the virus-specific DNA-binding proteins are structural proteins. Twelve are late proteins, the remaining being early proteins synthesized before viral DNA replication. Most of the virus-specific DNA-binding proteins bind both to double-stranded and to single-stranded DNA. The 110K, 29K, and 18K DNA-binding proteins bind only to single-stranded DNA. Two virus-specific enzymatic activities, DNA polymerase and RNA polymerase, were present in the fractions separated by DNA-cellulose chromatography. The virus-specific single-stranded DNA nuclease did not bind to DNA.
...
PMID:DNA-binding proteins specified by African swine fever virus. 368 26

We used UV light-induced cross-linking to study the interactions of cap binding proteins with the 5' cap structure of eucaryotic mRNAs. Thymidine kinase gene (herpes simplex virus type 1) transcripts prepared in vitro using the SP6 RNA polymerase transcription system were capped and methylated posttranscriptionally with [alpha-32P]GTP and S-adenosyl-L-methionine to yield cap-labeled transcripts. Irradiation of capped transcripts with crude rabbit reticulocyte initiation factors in the presence of ATP-Mg2+ resulted in the cap-specific cross-linking of two polypeptides with molecular masses of 24 and 80 kilodaltons (kDa). The cross-linking characteristics of these polypeptides resemble those of the cap-binding proteins previously detected by a chemical cross-linking assay (N. Sonenberg, D. Guertin, D. Cleveland, and H. Trachsel, Cell 27:563-572, 1981). However, the relative efficiency of the cross-linking of these two polypeptides to the cap structure was different from that in previous studies, and there was no detectable cross-linking of the previously described 50-kDa polypeptide. In addition, we present data indicating that the insertion of secondary structure into the 5' noncoding region of tk mRNA, 6 nucleotides from the cap structure, decreases the cap-specific cross-linking of the 80-kDa but not the 24-kDa polypeptide. In contrast, the insertion of secondary structure 37 nucleotides from the cap structure had no significant effect on the cross-linking of either the 24- or the 80-kDa cap-specific polypeptide. These results demonstrate that the position of mRNA 5'-proximal secondary structure relative to the cap structure can influence the cap-specific interaction between the mRNA and a translation initiation factor.
...
PMID:Photochemical cross-linking of cap binding proteins to eucaryotic mRNAs: effect of mRNA 5' secondary structure. 383 42

Short interspersed repetitive DNA sequences (SINEs) are the major component of dispersed repetitive DNA in all mammalian genomes. Most SINEs contain an intragenic RNA polymerase III promoter that initiates transcription at the 5' end of the repeated DNA sequence and which has been proposed to facilitate the transposition and amplification of these sequences by an RNA-intermediate mechanism. We have discovered several SINE families in the prosimian Galago crassicaudatus which have promoter regions similar to transfer RNA genes. To determine the relationship between Galago SINEs and mammalian tRNA genes, we have compared their sequences. Here, we demonstrate that the Galago monomer and type II SINE families are 68 and 62% homologous, respectively, with a human methionine tRNA gene. We have extended our analysis to include the rat identifier and mouse B2 families and show that their sequences are closely related to alanine and serine tRNA genes, respectively. Our observations suggest that many mammalian SINE families are amplified tRNA pseudogenes.
...
PMID:Repeat sequence families derived from mammalian tRNA genes. 385 Nov 63


<< Previous 1 2 3 4 5 6 7 8 9 10 Next >>

---