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. 2015 Oct 20;109(8):1537-40.
doi: 10.1016/j.bpj.2015.08.027.

Mapping the Processivity Determinants of the Kinesin-3 Motor Domain

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Free PMC article

Mapping the Processivity Determinants of the Kinesin-3 Motor Domain

Guido Scarabelli et al. Biophys J. .
Free PMC article

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 different cellular roles remains largely unknown. Here, we employed 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. We then used experimental mutagenesis and single-molecule motility assays to further assess the predicted residue-wise determinants of distinct kinesin-microtubule binding properties. Collectively, our results identify residues in the L8, L11, and α6 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.

Figures

Figure 1
Figure 1
Altering select family-specific tubulin interactions of kinesin-3 motors reduces processivity. (A) Refined molecular structures for kinesin-1 (green) and kinesin-3 (blue) resulting from cryo-electron microscopy (7) and subsequent molecular-dynamics simulations (see Supporting Materials and Methods for full details). (B) Differences in the residue contribution to the binding energy for kinesin-1 (green) and kinesin-3 (blue) in the ATP state. These values were determined from four replicate 40-ns molecular-dynamics simulations and subsequent energetic calculations. Note that specific interactions of the L2, L7, L8, L11, and α6 regions are predicted to enhance the binding affinity of kinesin-3 in relation to kinesin-1. (C) Processivity measurements from single-molecule motility assays of wild-type kinesin-1 (green), wild-type kinesin-3 (blue), and kinesin-3 mutants (red). The average run length (RL) and number of observations (N) are noted.

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