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The E-Hook of Tubulin Interacts with Kinesin's Head to Increase Processivity and Speed

Stefan Lakämper ^*  and Edgar Meyhöfer ^*

^* Department of Mechanical Engineering, University of Michigan, Ann Arbor, Michigan 48109; and Adolf Butenandt Institut, Zellbiologie, Universität München, 80336 Munich, Germany

Correspondence: Address reprint requests to Edgar Meyhöfer, Dept. of Mechanical Engineering, University of Michigan, 2350 Hayward, Ann Arbor, MI 48109. Tel.: 734-647-7856; Fax: 734-615-6647; E-mail: meyhofer{at}umich.edu.

Kinesins are dimeric motor proteins that move processively along microtubules. It has been proposed that the processivity of conventional kinesins is increased by electrostatic interactions between the positively charged neck of the motor and the negatively charged C-terminus of tubulin (E-hook). In this report we challenge this anchoring hypothesis by studying the motility of a fast fungal kinesin from Neurospora crassa (NcKin). NcKin is highly processive despite lacking the positive charges in the neck. We present a detailed analysis of how proteolytic removal of the E-hook affects truncated monomeric and dimeric constructs of NcKin. Upon digestion we observe a strong reduction of the processivity and speed of dimeric motor constructs. Monomeric motors with truncated or no neck display the same reduction of microtubule gliding speed as dimeric constructs, suggesting that the E-hook interacts with the head only. The E-hook has no effect on the strongly bound states of NcKin as microtubule digestion does not alter the stall forces produced by single dimeric motors, suggesting that the E-hook affects the interaction site of the kinesin·ADP-head and the microtubule. In fact, kinetic and binding experiments indicate that removal of the E-hook shifts the binding equilibrium of the weakly attached kinesin·ADP-head toward a more strongly bound state, which may explain reduced processivity and speed on digested microtubules.

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