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

The structure of the cAMP-CRP-CytR repression complex and the cAMP CRP-RNA polymerase initiation complex at the deoP2 promoter of E. coli have been probed by DNase I and uranyl footprinting. In the CRP2-CytR complex all protein DNA-phosphate contacts at CRP-1 and CRP-2 are retained, and in addition two new minor groove contacts, ascribed to phosphate-CytR interactions, are observed at -60 between the CRP sites. The contacts are compatible with a model in which the promoter DNA is wrapped around a complex of two CRPs and one CytR. In the RNA polymerase-CRP complex, the CRP-1 phosphate contacts are almost identical to those seen in the repression complex and strong RNA polymerase contacts are seen in the -10 and in the +10 regions. Most noteworthy are minor groove contacts in the -60 region ascribed to RNA polymerase contacts upstream from the CRP. Furthermore, binding of CRP to the CRP-2 target does not seem to interfere with RNA polymerase binding. Thus, a model is suggested in which the DNA is wrapped around a complex of RNA polymerase and one CRP. Finally, the results show that CytR and RNA polymerase are rivals that compete for binding with CRP at deoP2 and that CytR functions as an antiactivator.
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PMID:Characterization of promoter recognition complexes formed by CRP and CytR for repression and by CRP and RNA polymerase for activation of transcription on the Escherichia coli deoP2 promoter. 839 45

Activation of the two divergent Escherichia coli cai and fix operons involved in anaerobic carnitine metabolism is co-dependent on the cyclic AMP receptor protein (CRP) and on CaiF, the specific carnitine-sensitive transcriptional regulator. CaiF was overproduced using a phage T7 system, purified on a heparin column and ran as a 15 kDa protein on SDS-PAGE. DNase I footprinting and interference experiments identified two sites, F1 and F2, with apparently comparable affinities for the binding of CaiF in the cai-fix regulatory region. These sites share a common perfect inverted repeat comprising two 11 bp half-sites separated by 13 bp, and centred at -70 and -127 from the fix transcription start site. They were found to overlap the two low-affinity binding sites, CRP2 and CRP3, determined previously for CRP. Gel shift assays and footprinting experiments suggest that CaiF and CRP bind co-operatively to the F1/CRP2 and F2/CRP3 sites of the intergenic cai-fix region. Moreover, they appeared to serve the simultaneous binding of each other, giving rise to an original multiprotein CRP-CaiF complex enabling RNA polymerase recruitment and local DNA untwisting, at least at the fix promoter. Using random mutagenesis, two CaiF mutants impaired in transcription activation were isolated. The N-terminal A27V mutation affected the structural organization of the activator, whereas the central I62N mutation was suggested to interfere with DNA binding.
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PMID:Positive co-regulation of the Escherichia coli carnitine pathway cai and fix operons by CRP and the CaiF activator. 1056 97

Site-directed mutagenesis was conducted in the regulatory region of the Escherichia coli udp gene at promoter sites responsible for binding regulatory proteins CRP and CytR as well as RNA polymerase (the core-promoter containing the--10 sequence). In mutants with an "improved"--10 region, a partial relief from the control of the cAMP-CRP transcription activation complex occurred, and the negative CytR repressor regulation was reduced. In contrast, mutant promoters with a weak Pribnow block or with a deletion that completely eliminates the core-promoter exhibited an increased ability to titrate the CytR protein in vivo. On the other hand, the affinity of CytR for DNA in mutants with an altered--10 region was the same as in the wild-type udp promoter. After introduction of mutations affecting binding sites for CRP (CRP1 and CRP2), the negative effect of the CytR protein on promoter transcription was fully abolished. The CRP1 binding site was shown to play the main role in the activation of the promoter by the cAMP-CRP complex, whereas the CRP2 site participates in the formation of the repressor complex. Mutations in the main and additional CytR binding sites were isolated and characterized. On the basis of these data, it is concluded that the modification of each structural element of the udp regulatory region (binding sites for CytR, CRP, or RNA polymerase) caused changes in the overall pattern of the promoter regulation.
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PMID:[Functional interrelationship between elements of the Escherichia coli udp gene promotor responsible for binding regulatory proteins CytR, CRP, and RNA polymerase]. 1239 83

Effect of mutations in the -10 and -35 regions of the udp gene promoter on the nature of its regulation by CytR and CRP proteins was studied. In studies of expression of mutant promoters, competition between RNA polymerase and the CytR repressor for the promoter region of the udp gene was shown. In the presence of the improved -10 region, the introduction of a substitution 15C-->T (that is the presence of the elongated Pribnow block) resulted in the CRP-independent transcription of the udp gene promoter. The binding site CRP2 was shown to be indispensable for the maximum promoter activation by the transcription-activating cAMP-CRP complex. Both positive (cAMP-CRP complex) and negative (CytR) regulation of the promoter was virtually fully abolished after the introduction of mutations leading to the creation of canonical sequences in -10 and -35 promoter regions.
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PMID:[Structural-functional analysis of the promoter region of Escherichia coli udp gene]. 1502 96