Mapping the processivity determinants of the Kinesin-3 motor domain

Abstract

Kinesin superfamily members play important roles in many diverse cellular processes including cell motility, cell division, intracellular transport and regulation of the microtubule cytoskeleton. How the properties of the family-defining motor domain of distinct kinesins are tailored to their differing cellular roles remains largely unknown. Here we employ molecular dynamics simulations coupled with energetic calculations to infer the family-specific interactions of kinesin-1 and kinesin-3 motor domains with microtubules in different nucleotide states. Experimental mutagenesis and single molecule motility assays are then used to further assess the predicted residue-wise determinants of distinct kinesin-microtubule binding properties. Collectively, our results identify residues in the L8, L11 and alpha6 regions that contribute to family-specific microtubule interactions and whose mutation affects motor-microtubule complex stability and processive motility (the ability of an individual motor to take multiple steps along its microtubule filament). In particular, substitutions of prominent kinesin-3 residues with those found in kinesin-1, namely R167S/H171D, K266D, and R346M, were found to decrease kinesin-3 processivity 10-fold and thus approach kinesin-1 levels.