Skip to main content
U.S. flag

An official website of the United States government

Official websites use .gov
A .gov website belongs to an official government organization in the United States.

Secure .gov websites use HTTPS
A lock ( ) or https:// means you’ve safely connected to the .gov website. Share sensitive information only on official, secure websites.

Effect of Mutations at the Monomer-Monomer Interface of cAMP Receptor Protein on Specific DNA Binding



Y. Shi, S L. Wang, Susan Krueger, Frederick P. Schwarz


To Determine the thermodynamic role of binding of an operon to cAMP receptor protein (CRP) in the activation of transcription, isothermal titration calorimetry measurements were performed on the binding of three 40-base pair DNA sequences to the cyclic nucleoside complexes of CRP and its mutants at 296 K. The three 40 base pair sequences consisted of a consenus DNA (conDNA) duplex derived from the CRP-binding site sequences of the operons activated by CRP and two DNA sequences based on the CRP-binding site sequences of the lac operon (lacDNA) and the gal operon operon (galDNA). The mutants of CRP consisted of a T127L mutant, a S128A mutant, and a mutant containing both mutations (CRP*) which not only alter the transcriptional activity of the CRP complexes but also are involved in the monomer-monomer interfacial interactions of the CRP dimer. The binding reactions of the DNA duplexes to the fully cNMP-ligated CRP-mutant complexes were endothermic with binding constants as high as 6.6 + 1.1 x 106 M-1 (conDNA-CRP(cAMP)2). ConDNA binding to the unligated T127L and CRP* mutants was observed as well as conDNA and lacDNA binding to CRP with cAMP bound to only one monomer. The reduction of the binding constants with increase in KCI concentration indicated the formation of two ion pairs for the cAMP-ligated CRP and S128A complexes and four ion pairs for the cAMP-ligated T127L and CRP* complexes. Reduction of the DNA binding constants upon substitution of D2O for H2O in the buffer, the large heat capacity changes, and the enthalpy-entropy compensation exhibited by the bindingreactions indicate the importance of dehydration in the binding reaction. Small angle neutron scattering measurements on the lacDNA-CRP(cAMP)2 complex in D2O/H2O mixtures show that the DNA is bent around the cAMP-ligated protein in solution.
Journal of Biological Chemistry


cAMP receptor protein, DNA, isothermal titration calorimetry, thermodynamics, water


Shi, Y. , Wang, S. , Krueger, S. and Schwarz, F. (1999), Effect of Mutations at the Monomer-Monomer Interface of cAMP Receptor Protein on Specific DNA Binding, Journal of Biological Chemistry (Accessed June 22, 2024)


If you have any questions about this publication or are having problems accessing it, please contact

Created March 12, 1999, Updated February 19, 2017