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.

The Structure of the T127L/S128A Mutant of cAMP Receptor Protein Facilitates Promoter Site Binding



S. Chu, M. Tordova, G L. Gilliland, I Gorshkova, Y. Shi, S. Wang, Frederick P. Schwarz


The x-ray crystal structure of the cAMP-ligated T127L/S128A double mutant of cAMP receptor protein (CRP) was determined to a resolution of 2.2 . Although this structure is close to that of the x-ray crystal structure of cAMP-ligated CRP with one subunit in the open form and one subunit in the closed form, a bound syn-cAMP is clearly observed in the closed subunit in a third binding site in the closed subunit in a third binding site in the C-terminal domain. In addition, water-mediated interactions replace the hydrogen bonding interactions between the N6 of anti-cAMP bound in the N-terminal domains of each subunit and the OH groups of the Thr127 and Ser128 residues in the C α-helix of wild type CRP. This replacement induces flexibility in the C α-helix at Ala128, which swings the C-terminal domain of the open subunit more towards the N-terminal domain in the T127L/S128A double mutant of CRP(CRP*) than is observed in the open subunit of cAMP-ligated CRP. Isothermal titration calorimetry measurements on the binding of cAMP to CRP* show that the binding mechanism changes from an exothermic independent two-site binding mechanism at pH 7.0 to an endothermic interacting two-site mechanism at pH 5.2, similar to that observed for CRP at both pH levels. Differential scanning calorimetry measurements exhibit a broadening of the thermal denaturation transition of CRP* relative to that of CRP at pH = 7.0, but similar to the multipeak transitions observed for cAMP-ligated CRP. These properties and the bound syn-cAMP ligand, which has only been previously observed in the DNA bound x-ray crystal structure of cAMP-ligated by Passner and Steitz, (1997) Proc. Natl. Acad. Sci., U.S.A.94, 2843-2847, imply that the cAMP-ligated CRP* structure is closer to the conformation of the allosterically-activated structure than cAMP-ligated CRP. This may be induced by the unique pivotal point at Ala128 and/or by the bound syn-cAMP in the hinge region of CRP*.
Journal of Biological Chemistry


CAMP receptor protein, catabolate activator protein, differential scanning calorimetry, isothermal titration calorimetry, x-ray crystallography


Chu, S. , Tordova, M. , Gilliland, G. , Gorshkova, I. , Shi, Y. , Wang, S. and Schwarz, F. (2001), The Structure of the T127L/S128A Mutant of cAMP Receptor Protein Facilitates Promoter Site Binding, Journal of Biological Chemistry (Accessed December 10, 2023)
Created April 5, 2001, Updated October 12, 2021