Decoding Allosteric Communication Pathways in Cyclophilin A with a Comparative Analysis of Perturbed Conformational Ensembles

Conformational dynamics plays the key role in allosteric regulation of enzymes. Despite numerous experimental and computational efforts, the mechanism of how dynamics couple enzymatic function is poorly understood. Here, we introduce a new approach to exploring the dynamics-function relationship combining computational mutagenesis, microsecond-long molecular dynamics simulations, and side-chain torsion angle analyses. We apply our approach to elucidate the allosteric mechanism in cyclophilin A (CypA), a peptidyl-prolyl cis–trans isomerase known to participate in diverse biological processes and be associated with many diseases including cancer. Multiple single mutations are performed in CypA at previously discovered hotspot residues distal from the active site, and residues displaying significant dynamical changes upon mutations are then identified. The mutation-responsive residues delineate three distinct pathways potentially mediating allosteric communications between distal sites: two pathways resemble the allosteric networks identified in a recent experimental study, whereas the third represents a novel pathway. A residue–residue contact analysis is also performed to complement the findings. Furthermore, a recently developed difference contact network analysis is employed to explain mutation-specific allosteric effects. Our results suggest that comparing multiple conformational ensembles generated under various mutational conditions is a powerful tool to gain novel insights into enzymatic functions that are difficult to obtain through examining a single system such as the wild-type. Our approach is easy to extend for other systems. The results can also be utilized to facilitate the design of potent therapeutics targeting CypA