Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.7.6 (RNA polymerase)
 34,946 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A T7 RNA polymerase expression system has been used for the efficient expression of the yeast RNA polymerase general transcription factor TFIID (TFIIDY), the TATA-box factor (previously called BTF1) in Escherichia coli. Expression of the gene was performed at 25 degrees C instead of 37 degrees C to increase the total amount of soluble TFIIDY. Soluble TFIIDY was purified in three chromatographic steps and was eluted from the final column, a heparin-5PW HPLC column, in two peaks at 0.38 M (peak I) and 0.42 M (peak II) KCl in which this protein was 52% and greater than 95% pure, respectively. The protein in both peaks was active in an in vitro transcription assay. However, while TFIIDY from peak II was essentially indistinguishable from the material isolated from yeast, the protein of peak I differed in a number of biochemical characteristics, having a lower specific activity in an in vitro transcription assay and displaying an altered pattern of bands in a DNA band shift assay. Despite these differences, the proteins in both peaks have identical molecular weights on sodium dodecyl sulfate-polyacrylamide gel electrophoresis, have indistinguishable N-terminal amino acid sequences, and apparently exist as monomers under the conditions used for the heparin-5PW chromatography.
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PMID:Expression in Escherichia coli: purification and properties of the yeast general transcription factor TFIID. 182 18

A general transcription factor (BTF2) has been purified from HeLa whole cell extract and shown to be absolutely required for the formation of a functional initiation complex. We also demonstrate that this factor binds in solution to RNA polymerase B(II) and to the other general transcription factors BTF1(TFIID), BTF3, and BTF4. These results strongly suggest the existence of multiple interactions between the various components of the multiprotein initiation complex.
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PMID:Purification and interaction properties of the human RNA polymerase B(II) general transcription factor BTF2. 193 43

Fractionation of transcription extracts has led to the identification of multiple transcription factors specific for each form of nuclear RNA polymerase. Accurate transcription in vitro of the yeast U6 RNA gene by RNA polymerase C requires at least two factors. One of them was physically and functionally indistinguishable from transcription factor IID (TFIID or BTF1), a pivotal component of polymerase B transcription complexes, which binds to the TATA element. Purified yeast TFIID (yIID) or bacterial extracts that contained recombinant yIID were equally competent to direct specific transcription of the U6 gene by RNA polymerase C. The results suggest the formation of a hybrid transcription machinery, which may imply an evolutionary relation between class B and class C transcription factors.
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PMID:Participation of the TATA factor in transcription of the yeast U6 gene by RNA polymerase C. 198 75

We have fractionated rat liver and identified a set of transcription factors that are essential for accurate initiation by RNA polymerase II. These factors were resolved into five distinct enzyme fractions designated alpha, beta gamma, delta, epsilon, and tau. Four of these fractions can now be replaced with purified proteins. alpha and beta gamma were previously purified to apparent homogeneity (Conaway, J. W., and Conaway, R. C. (1989) J. Biol. Chem. 264, 2357-2362). Here, we report purification to near homogeneity of transcription factor epsilon. Epsilon has a native molecular mass of approximately 90 kDa and is composed of 34- and 58-kDa polypeptides. Both the 34- and 58-kDa polypeptides are required for runoff transcription. In addition, we show that transcription factor tau is a rat liver homologue of the TATA factor (TFIID or BTF1) that can be efficiently replaced in transcription in vitro by recombinant yeast TFIID. Comparison of the two factors reveals, however, that they differ significantly in their abilities to direct the transcription system to discriminate between promoters of different sequences.
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PMID:Transcription initiated by RNA polymerase II and transcription factors from liver. Structure and action of transcription factors epsilon and tau. 201 3

The initiation of transcription of eukaryotic genes involves the ordered assembly of a multiprotein complex on proximal promoter elements such as the TATA box. In addition to RNA polymerase II (otherwise RNA pol II, RNA polymerase B), four general transcription factors are required for initiation of transcription: BTF1 (also referred to as TFIID) which has recently been cloned from yeast, BTF2, BTF3 and STF. The first step in assembly of the initiation complex is the stable binding of BTF1 to the TATA box, which is facilitated by STF. Neither BTF2 nor BTF3 bind directly to the promoter proximal elements, but BTF3 can form a stable complex with RNA pol II. We recently purified BTF3, which is a protein of relative molecular mass 27,000, but further studies have been hampered by its low abundance in cells. On the basis of sequences from peptides of BTF3, we have now cloned two complementary DNAs, one for a protein (BTF3a) with all the characteristics of purified BTF3, and one for a shorter protein (BTF3b) lacking the first 44 residues of BTF3a and which is transcriptionally inactive, despite its ability to bind RNA pol II.
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PMID:Sequencing and expression of complementary DNA for the general transcription factor BTF3. 232 Jan 28

An activity (designated BTF1Y) in extracts of Saccharomyces cerevisiae can substitute for the human TATA box-binding factor BTF1 in a reconstituted transcription system containing the adenovirus 2 major late promoter, RNA polymerase B (II), and the basic transcription factors BTF2, BTF3, and STF. We have purified BTF1Y to homogeneity, using as assays reconstitution of in vitro transcription and DNase I footprinting on the TATA element. Both activities copurified with a 27-kDa polypeptide as determined by SDS/PAGE. Gel filtration indicated a molecular mass of 28 +/- 5 kDa under nondenaturing conditions, suggesting that the native BTF1Y protein is a monomer. BTF1Y was enzymatically cleaved, several peptides were sequenced, and appropriate oligonucleotide probes were synthesized to clone the BTF1Y gene from a yeast genomic library. The BTF1Y gene contains a 720-base-pair open reading frame encoding a protein of 27,003 Da. The recombinant protein expressed in HeLa cells exhibited the same chromatographic characteristics and in vitro transcriptional activity as BTF1Y prepared from yeast extracts, confirming the identity of the gene. Gene-disruption experiments indicated that the yeast BTF1Y gene is a single-copy essential gene.
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PMID:Cloning of the gene encoding the yeast protein BTF1Y, which can substitute for the human TATA box-binding factor. 269 73

Most class B (II) promoter regions from higher eukaryotes contain the TATA box and upstream and enhancer elements. Both the upstream and enhancer elements and their cognate factors have regulatory functions, whereas the TATA sequence interacts with the TATA box factor BTF1 to position RNA polymerase B and its ancillary initiation factors (STF, BTF2 and BTF3) to direct the initiation of transcription approximately 30 base pairs downstream. In many respects, class B promoter regions from the unicellular eukaryote Saccharomyces cerevisiae are similarly organized, containing upstream activating sequences that bear many similarities to enhancers. Although they are essential for initiation, the yeast TATA sequences are located at variable distances and further from the start sites (40-120 base pairs), whose locations are primarily determined by an initiator element. The basic molecular mechanisms that control initiation of transcription are known to be conserved from yeast to man: the yeast transcriptional transactivator GAL4 can activate a minimal TATA box-containing promoter in human HeLa cells, and a human inducible enhancer factor, the oestrogen receptor, can activate a similar minimal promoter in yeast. This striking evolutionary conservation prompted us to look for the presence in yeast of an activity that could possibly substitute for the human TATA box factor. We report here the existence of such an activity in yeast extracts.
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PMID:A yeast activity can substitute for the HeLa cell TATA box factor. 329 Jun 88