Dynamical network of residue–residue contacts reveals coupled allosteric effects in recognition, catalysis, and mutation

Significance How exactly protein motions facilitate substrate recognition and catalysis has remained largely unanswered. Characterization of protein dynamics at atomistic level is essential to understanding function. Molecular dynamics and NMR are helpful in this regard; however, analyzing multidimensional data from very long molecular dynamics (MD) simulations to elucidate key dynamical features observed in NMR remains very challenging. We present results from an approach for data analysis in which dynamics is defined in terms of interresidue contact formation and breaking. Analyzing simulation data on a therapeutically important Cyclophilin A and carrying out NMR experiments, we uncovered remarkable and unprecedented changes in its motions at a site over 15 Å from the active site upon substrate binding and how mutation in this distal site affects catalysis. Detailed understanding of how conformational dynamics orchestrates function in allosteric regulation of recognition and catalysis remains ambiguous. Here, we simulate CypA using multiple-microsecond-long atomistic molecular dynamics in explicit solvent and carry out NMR experiments. We analyze a large amount of time-dependent multidimensional data with a coarse-grained approach and map key dynamical features within individual macrostates by defining dynamics in terms of residue–residue contacts. The effects of substrate binding are observed to be largely sensed at a location over 15 Å from the active site, implying its importance in allostery. Using NMR experiments, we confirm that a dynamic cluster of residues in this distal region is directly coupled to the active site. Furthermore, the dynamical network of interresidue contacts is found to be coupled and temporally dispersed, ranging over 4 to 5 orders of magnitude. Finally, using network centrality measures we demonstrate the changes in the communication network, connectivity, and influence of CypA residues upon substrate binding, mutation, and during catalysis. We identify key residues that potentially act as a bottleneck in the communication flow through the distinct regions in CypA and, therefore, as targets for future mutational studies. Mapping these dynamical features and the coupling of dynamics to function has crucial ramifications in understanding allosteric regulation in enzymes and proteins, in general.