February 2010

[JBC] Kinesin’s Two-Water System

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Binding pocket of Eg5 kinesin with AMPPNP.

Many motor proteins like kinesins generate force and movement by coupling large-scale conformational changes to ATP binding and hydrolysis. However, a direct cause-and-effect linkage between ATP catalysis and powered structural changes in such proteins has not been firmly established, partially because the catalytic base that extracts a proton from the water nucleophile is unknown. In this study, the researchers determined the crystal structure of the motor domain of human Eg5 kinesin bound with the nonhydrolyzable ATP analogue AMPPNP, thus trapping the motor in a prehydrolytic state. They observed that, in this closed state, the two switch regions are linked by a salt bridge, and an ordered two-water cluster spans the distance between the inter-switch salt bridge and the AMPPNP γ-phosphate. This arrangement suggests that the second water molecule serves as a general base and shares a proton with the lytic water; the sequential transfer of a proton across this water network would disrupt the inter-switch salt-bridge, thereby promoting conformational transitions. This study provides the first experimental detection of the catalytic base for an ATPase and provides a mechanism that may apply to other NTPases with conserved active sites.

ATP Hydrolysis in Eg5 Kinesin Involves a Catalytic Two-water Mechanism 

Courtney L. Parke, Edward J. Wojcik, Sunyoung Kim and David K. Worthylake

J. Biol. Chem., published online Dec. 15, 2009 


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