Mapping the Structural and Dynamical Features of Kinesin Proteins

Abstract

Kinesin motor proteins drive intracellular transport by coupling ATP hydrolysis to conformational changes and directed movement along microtubules. Characterizing distinct conformations and their interconversion mechanism is thus essential to determining an atomic-level model of kinesin action. Here we report a comprehensive principal component analysis of 114 experimental structures along with the results of accelerated molecular dynamics simulations that together map the structural and dynamical features of the kinesin motor domain. All experimental structures were found to reside in one of eight distinct conformational clusters comprising two major groups. These groups differ in the orientation of key functional elements, including the microtubule binding alpha4-alpha5 subdomain. Group membership was found not to correlate with the nature of the bound nucleotide in a given structure. Accelerated molecular dynamics simulations of ATP, ADP and nucleotide free Eg5 indicated that all three nucleotide states could sample the major crystallographically observed conformations. Differences in the dynamic coupling of distal sites were evident in the simulations. In the ATP and APO simulations the neck-linker, loop8 and the alpha4-alpha5 subdomain display correlated motions that are absent in ADP simulations. Additional simulations predict that mutations G325A and G326A reduce the flexibility of these regions and disrupt their couplings. Furthermore, only in ADP simulations was the neck-linker region observed to undock. Additional APO simulations, commenced with an undocked neck-linker, formed coordinations reminiscent of the docked state. These interactions were absent in simulations of I359A mutants. Our combined results indicate that the reported ATP and ADP-like conformations of kinesin are intrinsically accessible regardless of nucleotide state. Furthermore, simulations highlight sites critical for large-scale conformational changes. We expect that further application of these methods will provide a framework for understanding the complete sequence of conformational changes and their relation to kinesin's ATPase cycle.