Structural snapshots of the kinesin‐2 OSM‐3 along its nucleotide cycle: implications for the ATP hydrolysis mechanism

Abstract

Motile kinesins are motor proteins that translocate along microtubules as they hydrolyze ATP. They share a conserved motor domain which harbors both ATPase and microtubule‐binding activities. An ATP hydrolysis mechanism involving two water molecules has been proposed based on the structure of the kinesin‐5 Eg5 bound to an ATP analog. Whether this mechanism is general in the kinesin superfamily remains uncertain. Here, we present structural snapshots of the motor domain of OSM‐3 along its nucleotide cycle. OSM‐3 belongs to the homodimeric kinesin‐2 subfamily and is the Caenorhabditis elegans homologue of human KIF17. OSM‐3 bound to ADP or devoid of a nucleotide shows features of ADP‐kinesins with a docked neck linker. When bound to an ATP analog, OSM‐3 adopts a conformation similar to those of several ATP‐like kinesins, either isolated or bound to tubulin. Moreover, the OSM‐3 nucleotide‐binding site is virtually identical to that of ATP‐like Eg5, demonstrating a shared ATPase mechanism. Therefore, our data extend to kinesin‐2 the two‐water ATP hydrolysis mechanism and further suggest that it is universal within the kinesin superfamily. Protein Database entries https://doi.org/10.2210/pdb7A3Z/pdb, https://doi.org/10.2210/pdb7A40/pdb, https://doi.org/10.2210/pdb7A5E/pdb.

Neurons dispose of hyperactive kinesin into glial cells for clearance

“…By what mechanism does a second missense substitution within the kinesin motor domain ameliorate the defects induced by the initial mutation in the hinge region? The ATP-binding motifs within the OSM-3 motor domain, akin to other extensively studied kinesin motors, encompass the P-loop, the phosphate-sensing loop known as Switch 1, and Switch 2 (Sweeney and Holzbaur, 2018 ; Varela et al, 2021 ). H207Q is situated in Switch 1 while R238W adjoins Switch 2 (Varela et al, 2021 ), with each substitution likely impeding nucleotide binding, thereby diminishing ATPase activity.…”

“…The ATP-binding motifs within the OSM-3 motor domain, akin to other extensively studied kinesin motors, encompass the P-loop, the phosphate-sensing loop known as Switch 1, and Switch 2 (Sweeney and Holzbaur, 2018 ; Varela et al, 2021 ). H207Q is situated in Switch 1 while R238W adjoins Switch 2 (Varela et al, 2021 ), with each substitution likely impeding nucleotide binding, thereby diminishing ATPase activity. Through microtubule-stimulated ATPase activity assays, we established that H207Q and R238W markedly attenuated ATPase activity to 18% and 39% comparing to the wild-type OSM-3, respectively (Fig.…”

A kinesin-1 variant reveals motor-induced microtubule damage in cells

“…For kinesin-1 proteins, zippering of the coverstrand (b0) with the neck linker (b9-b10) to form a two-stranded CNB is a critical first step in generation of a power stroke [21,31]. CNB formation has been also observed structurally for members of the kinesin-2, kinesin-3, kinesin-5 and kinesin-6 families [27,[45][46][47][48][49][50] and is critical for kinesin-3 force generation [23].…”