Kinetic studies of cAMP-induced propagation of the allosteric signal in the cAMP receptor protein from Escherichia coli with the use of site-directed mutagenesis

2009 ◽  
Vol 44 (3) ◽  
pp. 262-270 ◽  
Author(s):  
Andrzej Górecki ◽  
Barbara Kępys ◽  
Piotr Bonarek ◽  
Zygmunt Wasylewski
Keyword(s):  
Escherichia Coli ◽  
Kinetic Studies ◽  
Site Directed Mutagenesis ◽  
Receptor Protein ◽  
Directed Mutagenesis ◽  
Camp Receptor Protein ◽  
Camp Receptor

scholarly journals Positive and Negative Transcriptional Regulation of the Escherichia coli Gluconate Regulon Gene gntTby GntR and the Cyclic AMP (cAMP)-cAMP Receptor Protein Complex

1998 ◽  
Vol 180 (7) ◽  
pp. 1777-1785 ◽  
Author(s):  
Norbert Peekhaus ◽  
T. Conway
Keyword(s):  
Escherichia Coli ◽  
Cyclic Amp ◽  
Catabolite Repression ◽  
Transcriptional Start Site ◽  
Negative Regulator ◽  
Site Directed Mutagenesis ◽  
Receptor Protein ◽  
Start Site ◽  
Camp Receptor Protein ◽  
Camp Receptor

ABSTRACT The gntT gene of Escherichia coli is specifically induced by gluconate and repressed via catabolite repression. Thus, gluconate is both an inducer and a repressor ofgntT expression since gluconate is a catabolite-repressing sugar. In a gntR deletion mutant, the expression of a chromosomal gntT::lacZ fusion is both high and constitutive, confirming that GntR is the negative regulator of gntT. Indeed, GntR binds to two consensus gnt operator sites; one overlaps the −10 region of the gntT promoter, and the other is centered at +120 with respect to the transcriptional start site. The binding of GntR to these sites was proven in vitro by gel redardation assays and in vivo by site-directed mutagenesis of the binding sites. Binding of GntR to the operators is eliminated by gluconate and also by 6-phosphogluconate at a 10-fold-higher concentration. Interestingly, when gntR deletion strains are grown in the presence of gluconate, there is a twofold decrease in gntTexpression which is independent of catabolite repression and binding of GntR to the operator sites. This novel response of gntRmutants to the inducer is termed ultrarepression. Transcription ofgntT is activated by binding of the cyclic AMP (cAMP)-cAMP receptor protein (CRP) complex to a CRP binding site positioned at −71 upstream of the gntT transcription start site.

scholarly journals Cyclic AMP (cAMP) Receptor Protein-cAMP Complex Regulates Heparosan Production in Escherichia coli Strain Nissle 1917

2015 ◽  
Vol 81 (22) ◽  
pp. 7687-7696 ◽  
Author(s):  
Huihui Yan ◽  
Feifei Bao ◽  
Liping Zhao ◽  
Yanying Yu ◽  
Jiaqin Tang ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Cyclic Amp ◽  
Carbon Sources ◽  
Coli Strain ◽  
Site Directed Mutagenesis ◽  
Receptor Protein ◽  
Upstream Region ◽  
Camp Receptor Protein ◽  
Content Type ◽  
Camp Receptor

ABSTRACTHeparosan serves as the starting carbon backbone for the chemoenzymatic synthesis of heparin, a widely used clinical anticoagulant drug. The availability of heparosan is a significant concern for the cost-effective synthesis of bioengineered heparin. The carbon source is known as the pivotal factor affecting heparosan production. However, the mechanism by which carbon sources control the biosynthesis of heparosan is unclear. In this study, we found that the biosynthesis of heparosan was influenced by different carbon sources. Glucose inhibits the biosynthesis of heparosan, while the addition of either fructose or mannose increases the yield of heparosan. Further study demonstrated that the cyclic AMP (cAMP)-cAMP receptor protein (CRP) complex binds to the upstream region of the region 3 promoter and stimulates the transcription of the gene cluster for heparosan biosynthesis. Site-directed mutagenesis of the CRP binding site abolished its capability of binding CRP and eliminated the stimulative effect on transcription.1H nuclear magnetic resonance (NMR) analysis was further performed to determine theEscherichia colistrain Nissle 1917 (EcN) heparosan structure and quantify extracellular heparosan production. Our results add to the understanding of the regulation of heparosan biosynthesis and may contribute to the study of other exopolysaccharide-producing strains.

scholarly journals Interactions between the Escherichia coli cAMP receptor protein and the C-terminal domain of the α subunit of RNA polymerase at Class I promoters

1999 ◽  
Vol 337 (3) ◽  
pp. 415-423 ◽  
Author(s):  
Emma C. LAW ◽  
Nigel J. SAVERY ◽  
Stephen J. W. BUSBY
Keyword(s):  
Escherichia Coli ◽  
Rna Polymerase ◽  
Transcription Initiation ◽  
Class I ◽  
Receptor Protein ◽  
Camp Receptor Protein ◽  
Α Subunit ◽  
Terminal Domain ◽  
Up Elements ◽  
Camp Receptor

The Escherichia coli cAMP receptor protein (CRP) is a factor that activates transcription at over 100 target promoters. At Class I CRP-dependent promoters, CRP binds immediately upstream of RNA polymerase and activates transcription by making direct contacts with the C-terminal domain of the RNA polymerase α subunit (αCTD). Since αCTD is also known to interact with DNA sequence elements (known as UP elements), we have constructed a series of semi-synthetic Class I CRP-dependent promoters, carrying both a consensus DNA-binding site for CRP and a UP element at different positions. We previously showed that, at these promoters, the CRP–αCTD interaction and the CRP–UP element interaction contribute independently and additively to transcription initiation. In this study, we show that the two halves of the UP element can function independently, and that, in the presence of the UP element, the best location for the DNA site for CRP is position -69.5. This suggests that, at Class I CRP-dependent promoters where the DNA site for CRP is located at position -61.5, the two αCTDs of RNA polymerase are not optimally positioned. Two experiments to test this hypothesis are presented.

scholarly journals The cyclic AMP (cAMP)-cAMP receptor protein complex functions both as an activator and as a corepressor at the tsx-p2 promoter of Escherichia coli K-12.

1991 ◽  
Vol 173 (17) ◽  
pp. 5419-5430 ◽  
Author(s):  
P Gerlach ◽  
L Søgaard-Andersen ◽  
H Pedersen ◽  
J Martinussen ◽  
P Valentin-Hansen ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Cyclic Amp ◽  
Protein Complex ◽  
Receptor Protein ◽  
Camp Receptor Protein ◽  
Complex Functions ◽  
P2 Promoter ◽  
Camp Receptor ◽  
K 12

scholarly journals Structural Energy Landscapes and Plasticity of the Microstates of Apo Escherichia coli cAMP Receptor Protein

Biochemistry ◽  
2019 ◽  
Vol 59 (4) ◽  
pp. 460-470 ◽  
Author(s):  
Rati Chkheidze ◽  
Wilfredo Evangelista ◽  
Mark A. White ◽  
Y. Whitney Yin ◽  
J. Ching Lee
Keyword(s):  
Escherichia Coli ◽  
Receptor Protein ◽  
Energy Landscapes ◽  
Camp Receptor Protein ◽  
Camp Receptor

Determination of the rates of synthesis and degradation of adenosine 3′,5′-cyclic monophosphate in Escherichia coli CRP− and CRP+ strains

1978 ◽  
Vol 56 (9) ◽  
pp. 849-852 ◽  
Author(s):  
Ann D. E. Fraser ◽  
Hiroshi Yamazaki
Keyword(s):  
Escherichia Coli ◽  
Receptor Protein ◽  
Camp Receptor Protein ◽  
Balanced Growth ◽  
E Coli ◽  
Cyclic Monophosphate ◽  
Camp Receptor ◽  
Extracellular Camp ◽  
Synthesis And Degradation

We have developed a method for estimating the rates of synthesis and degradation of adenosine 3′,5′-cyclic monophosphate (cAMP) in Escherichia coli during balanced growth. Applying this method, we have found that an E. coli CRP− mutant 5333 (deficient for cAMP receptor protein) synthesizes cAMP about 25 times faster than does its CRP+ parent 1100. This accounts for the abnormally high intracellular and extracellular cAMP accumulation in 5333.

Sign in / Sign up

Export Citation Format

Share Document