Disease relevance of T7p07

• We have identified mutants of bacteriophage T7 RNA polymerase (RNAP) that are altered in their ability to pause or terminate at a variety of signals [1].
• The effect of phage T7 lysozyme on the production of biologically active porcine somatotropin in Escherichia coli from a gene transcribed by T7 RNA polymerase [2].
• The 31-kilodalton domain is composed of fingers, palm, and thumb subdomains arranged to form a DNA binding channel reminiscent of the polymerase domains of the Klenow fragment of Escherichia coli DNA polymerase I, HIV-1 reverse transcriptase, and bacteriophage T7 RNA polymerase [3].
• The recombinant vaccinia virus retained infectivity and stably expressed T7 RNA polymerase in mammalian cells [4].
• A T7 RNA polymerase chimeric gene containing a cauliflower mosaic virus 35S promoter and a tobacco ribulose-bisphosphate carboxylase/oxygenase small-subunit chloroplast transit-peptide sequence was introduced into tobacco by nuclear transformation [5].

High impact information on T7p07

• New crystal structures of transcription complexes formed by bacteriophage T7 RNA polymerase reveal a nucleotide-addition cycle driven by active-site conformational changes similar to those observed in DNA polymerases, and suggest provocative hypotheses for the more complex multisubunit RNA polymerases of free-living organisms [6].
• The structures of phage T7 RNA polymerase in an elongation phase substrate complex that includes the incoming nucleoside triphosphate and a pretranslocation product complex that includes the product pyrophosphate (PPi) are described here [7].
• The structural mechanism of translocation and helicase activity in T7 RNA polymerase [7].
• T7 lysozyme inhibits transcription by T7 RNA polymerase [8].
• Distinguishing between mechanisms of eukaryotic transcriptional activation with bacteriophage T7 RNA polymerase [9].

Chemical compound and disease context of T7p07

• Mature tRNA(Phe) containing a 5'-thiophosphate was synthesized by transcription in vitro using phage T7 RNA polymerase in the presence of guanosine 5'-phosphorothioate [10].
• The rat liver bifunctional enzyme, 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase (ATP:D-fructose-6-phosphate 2-phosphotransferase/D-fructose-2,6-bisphosphate 2-phosphohydrolase, EC 2.7.1.105/EC 3.1.3.46) and its separate kinase domain were expressed in Escherichia coli by using an expression system based on bacteriophage T7 RNA polymerase [11].
• The complex of T7 RNA polymerase with the phage phi 10 promoter has been visualized indirectly by exploiting the ability of the polymerase to protect DNA sequences from cleavage by methidiumpropyl-EDTA X Fe(II) [12].
• The kinetics of promoter binding and open complex formation in bacteriophage T7 RNA polymerase was investigated using 2-aminopurine (2-AP) modified promoters [13].
• Consistent with base-specific stabilizing interactions, of the 17 T7 RNA polymerase promoters in the phage genome, 15 begin with guanine [14].

Biological context of T7p07

• Higher levels of lysozyme supplied by plasmids pLysE or pLysH reduce the fully induced activity of T7 RNA polymerase such that induced cells can continue to grow and produce innocuous target proteins indefinitely [15].
• Nucleotide sequence of the gene for bacteriophage T7 RNA polymerase [16].
• However, the revisions do make a substantial difference in the amino acid sequence predicted for T7 RNA polymerase: 37 of the 883 amino acids are changed, 35 because of a shift in reading frame for one stretch of 37 amino acids [16].
• Lys631 residue in the active site of the bacteriophage T7 RNA polymerase. Affinity labeling and site-directed mutagenesis [17].
• The inserted DNA is a segment of 632 base pairs containing the complete coding sequence of both T7 gene 3.5 and weak promoter phi 3.8 for bacteriophage T7 RNA polymerase [18].

Anatomical context of T7p07

• Sequence similarities between phage RNA polymerases and those from mitochondria and chloroplasts, when interpreted in the context of our revised model of T7 RNA polymerase, suggest a conserved fold [19].
• The lysozyme of bacteriophage T7 is a bifunctional protein that cuts amide bonds in the bacterial cell wall and binds to and inhibits transcription by T7 RNA polymerase [20].
• T7 RNA polymerase-directed expression of an antibody fragment transgene in plastids causes a semi-lethal pale-green seedling phenotype [21].
• Here we show that transcription by phage T7 RNA polymerase from a divergent promoter can partially replace the requirement for circular Xenopus ribosomal RNA transcription templates in Xenopus oocytes [22].
• On the other hand, helper cells derived from the human embryonic kidney 293 cell line were generated constitutively expressing T7 RNA polymerase together with MV nucleocapsid protein and phosphoprotein [23].

Associations of T7p07 with chemical compounds

• The T7 RNA polymerase-nuclease is inhibited by T7 lysozyme and heparin, although not completely [24].
• After induction of T7 RNA polymerase by isopropyl beta-D-thiogalactopyranoside, a 16.5-kDa peptide accumulated to high levels [25].
• Equilibrium and stopped-flow kinetic studies of interaction between T7 RNA polymerase and its promoters measured by protein and 2-aminopurine fluorescence changes [26].
• Here, we have employed this system using a TAA ochre codon reversion assay to examine the fidelity of transcription by T7 RNA polymerase past an adenine residue adducted at the N6-position with (-)-anti-trans- or (+)-anti-trans-benzo[a]pyrene diol epoxide (BPDE) [27].
• It is proposed that T7 RNA polymerase interacts directly with the Hoogsteen side of the initial priming GTP (most likely via an interaction with an arginine side chain in the protein) to provide the extra stability required at this unique step in transcription [14].

Physical interactions of T7p07

• Structure of T7 RNA polymerase complexed to the transcriptional inhibitor T7 lysozyme [19].

Other interactions of T7p07

• Low levels of T7 lysozyme supplied by plasmids pLysS or pLysL, which are compatible with the pET vectors for expressing genes from a T7 promoter, are sufficient to stabilize many target plasmids and yet allow high levels of target protein to be produced upon induction of T7 RNA polymerase [15].
• Initiation of DNA synthesis can be reconstituted using T7 RNA polymerase, T7 DNA polymerase, and T7 origin-containing plasmid DNAs [28].

Analytical, diagnostic and therapeutic context of T7p07

• We have crystallized, using several approaches that may be of general interest, T7 RNA polymerase (T7RP) and the T7 RNA polymerase-T7 lysozyme complex (T7RPL) in forms suitable for structure determination by X-ray crystallography [29].
• HPLC gel-filtration experiments demonstrated complex formation between T7 lysozyme, T7 RNA polymerase, and promoter DNA [30].
• This overcomes the inhibitory effect of T7 lysozyme on T7 RNA polymerase as shown by SDS PAGE and flow cytometry analysis of expressed GFP [31].
• Genetic crosses demonstrated that a silent T7/GUS reporter gene could be activated in the F1 generation by transmission of an active nuclear T7 RNA polymerase gene from the male parent [5].
• As determined by indirect immunofluorescence, T7 RNA polymerase lacking the NLS remained mostly in the cytoplasm, whereas the protein containing the NLS localized to the nucleus [32].

References