Abstract
Allostery, the tranfer of information between distant parts of a macromolecule, is a fundamental feature of protein function and regulation. However, allosteric mechanisms are usually not explained by protein structure, requiring information on correlated fluctuations uniquely accessible to molecular simulation. Existing work to extract allosteric pathways from molecular dynamics simulations has focused on thermodynamic correlations. Here we show how kinetic correlations (i.e. dynamical activity) encode complementary information essential to explain observed variations in allosteric regulation. We performed atomistic simulations, totalling 0.5 milliseconds, on H, K, and NRas isoforms in the apo, GTP, and GDP-bound states of Ras protein, with and without complexing to its downstream effector, Raf. We show that differences in experimentally measured intrinsic and Raf-dependent catalytic speed amongst the three isoforms can be explained by dynamical activity and entropy, respectively. We show that Switch I and Switch II are the primary components of thermodynamic and kinetic allosteric networks, consistent with the key roles of these two motifs. These communication networks are altered by the hydrolysis of the substrate gamma phosphate, leading to increased entropy in HRas loops involved in substrate release. We find that the putative allosteric region is not coupled in KRas, but is coupled to the hydrolysis arm switch II in NRas and HRas, and that the mechanism in the latter two isoforms are thermodynamic and kinetic, respectively. Binding of Raf-RBD further activates thermodynamic allostery in HRas and KRas but has limited effect on NRas. These results indicate that kinetic and thermodynamic correlations are both needed to explain protein function and allostery. These two distinct channels of allosteric regulation, and their combinatorial variability, may explain how subtle mutational differences can lead to diverse regulatory profiles among enzymatic proteins.
Competing Interest Statement
The authors have declared no competing interest.
