Global Protein Motions in the Allosteric Regulation of CRP/FNR Family Transcription Factors

Allostery is a fundamental process by which ligand binding to a protein alters its activity at a distinct site. There is growing evidence that allosteric co-operativity can be communicated by modulation of protein dynamics. We provide a foundational theory for how allostery can occur as a function of low frequency dynamics without a change in protein structure. We have generated coarse-grained models that describe the protein backbone motions of the homodimeric CRP/FNR family transcription factors, Catabolite Activated Protein (CAP) and GlxR. We demonstrate that binding the first molecule of cAMP ligand modulates the global normal modes resulting in negative co-operativity for binding the second cAMP ligand without a change in mean structure. The theory makes key experimental predictions and these are validated through an analysis of variant proteins by a combination of structural biology and isothermal calorimetry. A quantitative description of allostery as a free energy landscape revealed a protein ‘design space’ that identified the key inter- and intramolecular regulatory parameters that contribute to CRP/FNR family allostery.