Influence of the Location of the cAMP Receptor Protein Binding Site on the Geometry of a Transcriptional Activation Complex inEscherichiacoli†

Biochemistry ◽  
1996 ◽  
Vol 35 (48) ◽  
pp. 15302-15312 ◽  
Author(s):  
Patrick Eichenberger ◽  
Sylvie Déthiollaz ◽  
Nobuyuki Fujita ◽  
Akira Ishihama ◽  
Johannes Geiselmann
Keyword(s):  
Protein Binding ◽  
Binding Site ◽  
Transcriptional Activation ◽  
Receptor Protein ◽  
Protein Binding Site ◽  
Camp Receptor Protein ◽  
Camp Receptor

scholarly journals Functional homology between the yeast regulatory proteins GAL4 and LAC9: LAC9-mediated transcriptional activation in Kluyveromyces lactis involves protein binding to a regulatory sequence homologous to the GAL4 protein-binding site.

1987 ◽  
Vol 7 (12) ◽  
pp. 4400-4406 ◽  
Author(s):  
K D Breunig ◽  
P Kuger
Keyword(s):  
Protein Binding ◽  
Binding Site ◽  
Transcriptional Activation ◽  
Kluyveromyces Lactis ◽  
Regulatory Protein ◽  
Binding Activity ◽  
Protein Binding Site ◽  
Regulatory Sequence ◽  
Functional Homology ◽  
Beta Galactosidase

As shown previously, the beta-galactosidase gene of Kluyveromyces lactis is transcriptionally regulated via an upstream activation site (UASL) which contains a sequence homologous to the GAL4 protein-binding site in Saccharomyces cerevisiae (M. Ruzzi, K.D. Breunig, A.G. Ficca, and C.P. Hollenberg, Mol. Cell. Biol. 7:991-997, 1987). Here we demonstrate that the region of homology specifically binds a K. lactis regulatory protein. The binding activity was detectable in protein extracts from wild-type cells enriched for DNA-binding proteins by heparin affinity chromatography. These extracts could be used directly for DNase I and exonuclease III protection experiments. A lac9 deletion strain, which fails to induce the beta-galactosidase gene, did not contain the binding factor. The homology of LAC9 protein with GAL4 (J.M. Salmeron and S. A. Johnston, Nucleic Acids Res. 14:7767-7781, 1986) strongly suggests that LAC9 protein binds directly to UASL and plays a role similar to that of GAL4 in regulating transcription.

scholarly journals Regulation of the expression of whiB1 in Mycobacterium tuberculosis: role of cAMP receptor protein

Microbiology ◽  
2006 ◽  
Vol 152 (9) ◽  
pp. 2749-2756 ◽  
Author(s):  
Nisheeth Agarwal ◽  
Tirumalai R. Raghunand ◽  
William R. Bishai
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Binding Site ◽  
Receptor Protein ◽  
Camp Receptor Protein ◽  
Mobility Shift ◽  
Camp Analogue ◽  
Camp Receptor ◽  
Fusion Analysis ◽  
Electrophoretic Mobility Shift Assays

The wbl (whiB-like) genes encode putative transcription factors unique to actinomycetes. This study characterized the promoter element of one of the seven wbl genes of Mycobacterium tuberculosis, whiB1 (Rv3219c). The results reveal that whiB1 is transcribed by a class I-type cAMP receptor protein (CRP)-dependent promoter, harbouring a CRP-binding site positioned at −58.5 with respect to its transcription start point. In vivo promoter activity analysis and electrophoretic mobility shift assays suggest that the expression of whiB1 is indeed regulated by cAMP-dependent binding of CRPM (encoded by the M. tuberculosis gene Rv3676) to the whiB1 5′ untranslated region (5′UTR). β-Galactosidase gene fusion analysis revealed induction of the whiB1 promoter in M. tuberculosis on addition of exogenous dibutyric cAMP (a diffusible cAMP analogue) only when an intact CRP-binding site was present. These results indicate that M. tuberculosis whiB1 transcription is regulated in part by cAMP levels via direct binding of cAMP-activated CRPM to a consensus CRP-binding site in the whiB1 5′UTR.

scholarly journals Mutations that affect transcription and cyclic AMP-CRP regulation of the adenylate cyclase gene (cya) of Salmonella typhimurium.

Genetics ◽  
1990 ◽  
Vol 125 (4) ◽  
pp. 719-727
Author(s):  
J P Fandl ◽  
L K Thorner ◽  
S W Artz
Keyword(s):  
Salmonella Typhimurium ◽  
Binding Site ◽  
Minimal Medium ◽  
Receptor Protein ◽  
Camp Receptor Protein ◽  
Multiple Promoters ◽  
P2 Promoter ◽  
Camp Receptor ◽  
Insertion Mutations

Abstract We studied the expression of the cya promoter(s) in cya-lac fusion strains of Salmonella typhimurium and demonstrated cAMP receptor protein (CRP)-dependent repression by cAMP. Expression of cya was reduced about fourfold in cultures grown in acetate minimal medium as compared to cultures grown in glucose-6-phosphate minimal medium. Expression of cya was also reduced about fourfold by addition of 5 mM cAMP to cultures grown in glucose minimal medium. We constructed in vitro deletion and insertion mutations altering a major cya promoter (P2) and a putative CRP binding site overlapping P2. These mutations were recombined into the chromosome by allele replacement with M13mp::cya recombinant phages and the regulation of the mutant promoters was analyzed. A 4-bp deletion of the CRP binding site and a 4-bp insertion in this site nearly eliminated repression by cAMP. A mutant with the P2 promoter and the CRP binding site both deleted exhibited an 80% reduction in cya expression; the 20% residual expression was insensitive to cAMP repression. This mutant retained a Cya+ phenotype. Taken together, the results establish that the cya gene is transcribed from multiple promoters one of which, P2, is negatively regulated by the cAMP-CRP complex. Correction for the contribution to transcription by the cAMP-CRP nonregulated cya promoters indicates that the P2 promoter is repressed at least eightfold by cAMP-CRP.

scholarly journals Spacing requirements for interactions between the C-terminal domain of the α subunit of Escherichia coli RNA polymerase and the cAMP receptor protein

1998 ◽  
Vol 330 (1) ◽  
pp. 413-420 ◽  
Author(s):  
S. Georgina LLOYD ◽  
J. W. Stephen BUSBY ◽  
J. Nigel SAVERY
Keyword(s):  
Escherichia Coli ◽  
Rna Polymerase ◽  
Binding Site ◽  
Transcription Initiation ◽  
Receptor Protein ◽  
Camp Receptor Protein ◽  
Α Subunit ◽  
Synthetic Promoters ◽  
Terminal Domain ◽  
Camp Receptor

During transcription initiation at bacterial promoters, the C-terminal domain of the RNA polymerase α subunit (αCTD) can interact with DNA-sequence elements (known as UP elements) and with activator proteins. We have constructed a series of semi-synthetic promoters carrying both an UP element and a consensus DNA-binding site for the Escherichia coli cAMP receptor protein (CRP; a factor that activates transcription by making direct contacts with αCTD). At these promoters, the UP element was located at a variety of distances upstream of the CRP-binding site, which was fixed at position -41.5 bp upstream of the transcript start. At some positions, the UP element caused enhanced promoter activity whereas, at other positions, it had very little effect. In no case was the CRP-dependence of the promoter relieved. DNase I and hydroxyl-radical footprinting were used to study ternary RNA polymerase-CRP-promoter complexes formed at two of the most active of these promoters, and co-operativity between the binding of CRP and purified α subunits was studied. The footprints show that αCTD binds to the UP element as it is displaced upstream but that this displacement does not prevent αCTD from being contacted by CRP. Models to account for this are discussed.

Functional homology between the yeast regulatory proteins GAL4 and LAC9: LAC9-mediated transcriptional activation in Kluyveromyces lactis involves protein binding to a regulatory sequence homologous to the GAL4 protein-binding site

1987 ◽  
Vol 7 (12) ◽  
pp. 4400-4406
Author(s):  
K D Breunig ◽  
P Kuger
Keyword(s):  
Protein Binding ◽  
Binding Site ◽  
Transcriptional Activation ◽  
Kluyveromyces Lactis ◽  
Regulatory Protein ◽  
Binding Activity ◽  
Protein Binding Site ◽  
Regulatory Sequence ◽  
Functional Homology ◽  
Beta Galactosidase

As shown previously, the beta-galactosidase gene of Kluyveromyces lactis is transcriptionally regulated via an upstream activation site (UASL) which contains a sequence homologous to the GAL4 protein-binding site in Saccharomyces cerevisiae (M. Ruzzi, K.D. Breunig, A.G. Ficca, and C.P. Hollenberg, Mol. Cell. Biol. 7:991-997, 1987). Here we demonstrate that the region of homology specifically binds a K. lactis regulatory protein. The binding activity was detectable in protein extracts from wild-type cells enriched for DNA-binding proteins by heparin affinity chromatography. These extracts could be used directly for DNase I and exonuclease III protection experiments. A lac9 deletion strain, which fails to induce the beta-galactosidase gene, did not contain the binding factor. The homology of LAC9 protein with GAL4 (J.M. Salmeron and S. A. Johnston, Nucleic Acids Res. 14:7767-7781, 1986) strongly suggests that LAC9 protein binds directly to UASL and plays a role similar to that of GAL4 in regulating transcription.

Sign in / Sign up

Export Citation Format

Share Document