title: The Role of Protein-Ligand Contacts in Allosteric Regulation of the Escherichia coli Catabolite Activator Protein.
abstract: Allostery is a fundamental process by which ligand binding to a protein alters its activity at a distant site. Both experimental and theoretical evidence demonstrate that allostery can be communicated through altered slow relaxation protein dynamics without conformational change. The catabolite activator protein (CAP) of Escherichia coli is an exemplar for the analysis of such entropically driven allostery. Negative allostery in CAP occurs between identical cAMP binding sites. Changes to the cAMP-binding pocket can therefore impact the allosteric properties of CAP. Here we demonstrate, through a combination of coarse-grained modeling, isothermal calorimetry, and structural analysis, that decreasing the affinity of CAP for cAMP enhances negative cooperativity through an entropic penalty for ligand binding. The use of variant cAMP ligands indicates the data are not explained by structural heterogeneity between protein mutants. We observe computationally that altered interaction strength between CAP and cAMP variously modifies the change in allosteric cooperativity due to second site CAP mutations. As the degree of correlated motion between the cAMP-contacting site and a second site on CAP increases, there is a tendency for computed double mutations at these sites to drive CAP toward noncooperativity. Naturally occurring pairs of covarying residues in CAP do not display this tendency, suggesting a selection pressure to fine tune allostery on changes to the CAP ligand-binding pocket without a drive to a noncooperative state. In general, we hypothesize an evolutionary selection pressure to retain slow relaxation dynamics-induced allostery in proteins in which evolution of the ligand-binding site is occurring.
authors.affiliation: From the School of Biological and Biomedical Sciences, the Biophysical Sciences Institute, and.
authors.forename: Philip D
authors.initials: PD
authors.lastname: Townsend
authors.affiliation: the School of Chemical Engineering and Analytical Science, University of Manchester, Manchester M13 9PL, United Kingdom.
authors.forename: Thomas L
authors.initials: TL
authors.lastname: Rodgers
authors.affiliation: From the School of Biological and Biomedical Sciences, the Biophysical Sciences Institute, and the Departments of Chemistry and.
authors.forename: Laura C
authors.initials: LC
authors.lastname: Glover
authors.affiliation: the Biophysical Sciences Institute, and the Departments of Chemistry and the Department of Chemistry, University of Turku, 20014 Turku, Finland, and.
authors.forename: Heidi J
authors.initials: HJ
authors.lastname: Korhonen
authors.affiliation: From the School of Biological and Biomedical Sciences.
authors.forename: Shane A
authors.initials: SA
authors.lastname: Richards
authors.affiliation: the Department of Chemistry, University of Cambridge, Cambridge CB2 1EW, United Kingdom.
authors.forename: Lucy J
authors.initials: LJ
authors.lastname: Colwell
authors.affiliation: From the School of Biological and Biomedical Sciences, the Biophysical Sciences Institute, and the Departments of Chemistry and.
authors.forename: Ehmke
authors.initials: E
authors.lastname: Pohl
authors.affiliation: the Biophysical Sciences Institute, and the Departments of Chemistry and.
authors.forename: Mark R
authors.initials: MR
authors.lastname: Wilson
authors.affiliation: the Biophysical Sciences Institute, and the Departments of Chemistry and.
authors.forename: David R W
authors.initials: DR
authors.lastname: Hodgson
authors.affiliation: the Biophysical Sciences Institute, and the Departments of Chemistry and Physics, Durham University, Durham DH1 3LE, United Kingdom.
authors.forename: Tom C B
authors.initials: TC
authors.lastname: McLeish
authors.affiliation: From the School of Biological and Biomedical Sciences, the Biophysical Sciences Institute, and m.j.cann@durham.ac.uk.
authors.forename: Martin J
authors.initials: MJ
authors.lastname: Cann
mesh_terms: D000494:Allosteric Regulation; D001665:Binding Sites; D018360:Crystallography, X-Ray; D000242:Cyclic AMP / Q000737:chemistry* / Q000378:metabolism; D002373:Cyclic AMP Receptor Protein / Q000737:chemistry* / Q000378:metabolism; D019277:Entropy; D004926:Escherichia coli / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000378:metabolism; D008024:Ligands; D008968:Molecular Conformation; D056004:Molecular Dynamics Simulation; D015394:Molecular Structure; D011485:Protein Binding; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary
chemical_list: D002373:Cyclic AMP Receptor Protein; D029968:Escherichia coli Proteins; D008024:Ligands; C481662:crp protein, E coli; D000242:Cyclic AMP
keywords: allosteric regulation; biophysics; calorimetry; cyclic AMP (cAMP); nucleoside/nucleotide analogue; protein dynamic