Binding sites on microtubules of kinesin motors of the same or opposite polarity.

Abstract

The kinesin motor proteins translocate toward either the plus or minus end of microtubules (MTs). Competitive microtubule binding assays were carried out with monomeric motor domains of the minus-end-directed nonclaret disjunctional (Ncd) and Kar3 and the plus-end-directed kinesin heavy chain (KHC) to determine whether motors of the same or opposite polarity compete for binding sites on MTs and to test the idea that motor polarity is determined by differences in binding sites on MTs of the motors. The stoichiometries of binding were approximately 1 motor:1 tubulin heterodimer for all three motors. Ncd and Kar3, both minus-end motors, severely inhibited the binding of one another to MTs, as predicted theoretically for binding of the two motors to the same site on MTs, indicating that the binding sites on MTs of Ncd and Kar3 are the same or overlap extensively. Motors of opposite polarity, KHC and Ncd or KHC and Kar3, showed partial or complete inhibition of binding to MTs under different experimental protocols. The differences in binding behavior could be due to experimental conditions or be inherent in the nature of motor binding to MTs. Alternatively, differences in KHC and Ncd or Kar3 binding sites on MTs may exist such that the motors bind to partially overlapping but nonidentical sites on MTs. These differences in binding sites may be related to the opposite polarity of translocation on MTs of the motors.