Allosteric Response of DNA Recognition Helices of Catabolite Activator Protein to cAMP and DNA Binding.

The homodimeric catabolite activator protein (CAP) regulates the transcription of several bacterial genes based on the cellular concentration of cyclic adenosine monophosphate (cAMP). The binding of cAMP to CAP triggers allosteric communication between the cAMP binding domains (CBD) and DNA binding domains (DBD) of CAP, which entails repositioning of DNA recognition helices (F-helices) in the DBD to dock favorably to the target DNA. Despite considerable progress, much remains to be understood about the mechanistic details of DNA recognition by CAP and about the map of allosteric pathways involved in CAP-mediated gene transcription. The present study uses molecular dynamics and umbrella sampling simulations to investigate the mechanism of cAMP- and DNA-induced changes in the conformation and energetics of F-helices observed during the allosteric regulation of CAP by cAMP and the subsequent binding to the DNA promoter region. Using novel collective variables, the free energy profiles associated with the orientation and dynamics of F-helices in the unliganded, cAMP-bound, and cAMP-DNA-bound states of CAP are calculated and compared. The binding-induced alterations in the resultant free energy profiles reveal important flexibility constraints imposed on DBD upon cAMP and DNA binding. A comprehensive analysis of residue-wise interaction maps reveals potential allosteric pathways between CBD and DBD that facilitate the allosteric transduction of regulatory signals in CAP. The revelation that the predicted allosteric pathways crisscross the intersubunit interface offers important clues on the microscopic origin of the intersubunit cooperativity and dimer stability of CAP.