Effect of Charge on Autoinhibition of Kinesin-1

Abstract

Kinesin-1 is autoinhibited at physiological salt concentrations through an inhibitory interaction of C-terminal tail domains with the N-terminal motor (head) domains. This interaction of heads with the tail is highly dependent on the ionic strength, with high ionic strength strongly favoring dissociation. Unexpectedly the binding stoichiometry is one tail peptide binding to the two heads of a motor domain dimer, as originally indicated by biochemical titrations and recently confirmed by the determination of the structure of a head dimer - tail complex by X-ray crystallography (Kaan, et al., Science 333, 832 (2011)). A short region of the tail centered on lysine-944 (the IAK region) is sufficient for binding of a single tail to two heads. However, the binding affinity of this short region on its own is low. Tight binding of the tail to the heads requires the inclusion of an adjacent region with a large excess of positive charge. This positive region is designated the auxiliary binding site (ABS) because it is also responsible for the nucleotide-independent binding to the tail to microtubules. Kinesin motor domains have several negatively charged side chains near where the heads are cross linked by the tail domain. Mutation of these negative head residues to Ala decreases the affinity for heads of tail domains that contain both the ABS and the IAK region. This suggests that the increased affinity produced by the ABS region is due to interactions with these negative regions of the heads.Supported by NIH grant NS058848.