scholarly journals Binding of the cyclic AMP receptor protein of Escherichia coli and DNA bending at the P4 promoter of pBR322

1992 ◽  
Vol 285 (1) ◽  
pp. 91-97 ◽  
Author(s):  
I Brierley ◽  
J G Hoggett
Keyword(s):  
Escherichia Coli ◽  
Cyclic Amp ◽  
Gel Electrophoresis ◽  
Dna Bending ◽  
Receptor Protein ◽  
Simple Relationship ◽  
Start Site ◽  
Transcription Start ◽  
Bending Angle ◽  
Cyclic Amp Receptor Protein

The binding of the Escherichia coli cyclic AMP receptor protein (CRP) to its specific site on the P4 promoter of pBR322 has been studied by gel electrophoresis. Binding to the P4 site was about 40-50-fold weaker than to the principal CRP site on the lactose promoter at both low (0.01 M) and high (0.1 M) ionic strengths. CRP-induced bending at the P4 site was investigated from the mobilities of CRP bound to circularly permuted P4 fragments. The estimated bending angle, based on comparison with Zinkel & Crothers [(1990) Biopolymers 29, 29-38] A-tract bending standards, was found to be approximately 96 degrees, similar to that found for binding to the lac site. These observations suggest that there is not a simple relationship between strength of CRP binding and the extent of induced bending for different CRP sites. The apparent centre of bending in P4 is displaced about 6-8 bp away from the conserved TGTGA sequence and the P4 transcription start site.

scholarly journals Repression by Cyclic AMP Receptor Protein at a Distance

mBio ◽  
2012 ◽  
Vol 3 (5) ◽  
Author(s):  
David J. Lee ◽  
Stephen J. W. Busby
Keyword(s):  
Escherichia Coli ◽  
Transcription Factors ◽  
Cyclic Amp ◽  
Promoter Activity ◽  
Bacterial Genome ◽  
Receptor Protein ◽  
Transcription Start ◽  
Content Type ◽  
Cyclic Amp Receptor Protein ◽  
Α Subunits

ABSTRACT In a previous study of promoters dependent on the Escherichia coli cyclic AMP receptor protein (CRP), carrying tandem DNA sites for CRP, we found that the upstream-bound CRP could either enhance or repress transcription, depending on its location. Here, we have analyzed the interactions between CRP and the C-terminal domains of the RNA polymerase α subunits at some of these promoters. We report that the upstream-bound CRP interacts with these domains irrespective of whether it up- or downregulates promoter activity. Hence, disruption of this interaction can lead to either down- or upregulation, depending on its location. IMPORTANCE Many bacterial promoters carry multiple DNA sites for transcription factors. While most factors that downregulate promoter activity bind to targets that overlap or are downstream of the transcription start and −10 element, very few cases of repression from upstream locations have been reported. Since more Escherichia coli promoters are regulated by cyclic AMP receptor protein (CRP) than by any other transcription factor, and since multiple DNA sites for CRP are commonplace at promoters, our results suggest that promoter downregulation by transcription factors may be more prevalent than hitherto thought, and this will have implications for the annotation of promoters from new bacterial genome sequences.

Mannose utilization in Escherichia coli requires cyclic AMP but not an exogenous inducer

1980 ◽  
Vol 26 (12) ◽  
pp. 1508-1511 ◽  
Author(s):  
Ann D. E. Fraser ◽  
Hiroshi Yamazaki
Keyword(s):  
Escherichia Coli ◽  
Carbon Source ◽  
Cyclic Amp ◽  
Adenylyl Cyclase ◽  
Sole Carbon Source ◽  
Receptor Protein ◽  
Deficient Mutant ◽  
Phosphotransferase System ◽  
Cyclic Amp Receptor Protein ◽  
Cyclic Monophosphate

It has not been clarified whether the utilization of mannose by Escherichia coli requires adenosine 3′,5′-cyclic monophosphate (cyclic AMP). Using an adenylyl cyclase deficient mutant (CA8306B) and a cyclic AMP receptor protein (CRP) deficient mutant (5333B) we have shown that the utilization of mannose is dependent on the cyclic AMP–CRP complex. 2-Deoxyglucose (DG) is a nonmetabolizable glucose analog specific for the phosphotransferase system (PTS) which transports mannose (termed here PTSM). Growth of CA8306B on glycerol is unaffected by addition of the analog, whereas growth of the strain on glycerol plus cyclic AMP ceases im mediately upon addition of DG. These results suggest that the formation of PTSM is dependent on cyclic AMP. In addition, CA8306B grown on glycerol plus cyclic AMP can immediately utilize mannose when transferred to a medium containing mannose as a sole carbon source, whereas the same strain grown on glycerol without cyclic AMP cannot utilize mannose when so transferred. These results suggest that the formation of PTSM does not require an exogenous inducer.

scholarly journals Mutations in the cyclic AMP binding site of the cyclic AMP receptor protein of Escherichia coli

1988 ◽  
Vol 253 (3) ◽  
pp. 801-807 ◽  
Author(s):  
A M Gronenborn ◽  
R Sandulache ◽  
S Gärtner ◽  
G M Clore
Keyword(s):  
Escherichia Coli ◽  
Cyclic Amp ◽  
Binding Site ◽  
Cyclic Nucleotides ◽  
Binding Pocket ◽  
Receptor Protein ◽  
Directed Mutagenesis ◽  
Wild Type ◽  
Cyclic Amp Receptor Protein ◽  
Better Than

Mutants in the cyclic AMP binding site of the cyclic AMP receptor protein (CRP) of Escherichia coli have been constructed by oligonucleotide-directed mutagenesis. They have been phenotypically characterized and their ability to enhance the expression of catabolite-repressible operons has been tested. In addition, the binding of cyclic nucleotides to the mutants has been investigated. It is shown that the six mutants made fall into one of three classes: (i) those that bind cyclic AMP better than the wild type protein (Ser-62→Ala) and result in greater transcription enhancement; (ii) those that bind cyclic AMP similarly to wild type (Ser-83→Ala, Ser-83→Lys, Thr-127→Ala, Ser-129→Ala); and (iii) those that do not bind cyclic AMP at all (Arg-82→Leu). Implications of these findings with respect to present models of the cyclic nucleotide binding pocket of CRP are discussed.

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