Kinesin-3 motors are fine-tuned at the molecular level to endow distinct mechanical outputs

Soppina, PushpanjaliPushpanjaliSoppinaPatel, NishabenNishabenPatelShewale, Dipeshwari J.Dipeshwari J.ShewaleRai, AshimAshimRaiSivaramakrishnan, SivarajSivarajSivaramakrishnanNaik, Pradeep K.Pradeep K.NaikSoppina, VirupakshiVirupakshiSoppina2025-08-312025-08-312022-12-0110.1186/s12915-022-01370-82-s2.0-85135601957https://d8.irins.org/handle/IITG2025/2583935948971Background: Kinesin-3 family motors drive diverse cellular processes and have significant clinical importance. The ATPase cycle is integral to the processive motility of kinesin motors to drive long-distance intracellular transport. Our previous work has demonstrated that kinesin-3 motors are fast and superprocessive with high microtubule affinity. However, chemomechanics of these motors remain poorly understood. Results: We purified kinesin-3 motors using the Sf9-baculovirus expression system and demonstrated that their motility properties are on par with the motors expressed in mammalian cells. Using biochemical analysis, we show for the first time that kinesin-3 motors exhibited high ATP turnover rates, which is 1.3- to threefold higher compared to the well-studied kinesin-1 motor. Remarkably, these ATPase rates correlate to their stepping rate, suggesting a tight coupling between chemical and mechanical cycles. Intriguingly, kinesin-3 velocities (KIF1A > KIF13A > KIF13B > KIF16B) show an inverse correlation with their microtubule-binding affinities (KIF1A < KIF13A < KIF13B < KIF16B). We demonstrate that this differential microtubule-binding affinity is largely contributed by the positively charged residues in loop8 of the kinesin-3 motor domain. Furthermore, microtubule gliding and cellular expression studies displayed significant microtubule bending that is influenced by the positively charged insert in the motor domain, K-loop, a hallmark of kinesin-3 family. Conclusions: Together, we propose that a fine balance between the rate of ATP hydrolysis and microtubule affinity endows kinesin-3 motors with distinct mechanical outputs. The K-loop, a positively charged insert in the loop12 of the kinesin-3 motor domain promotes microtubule bending, an interesting phenomenon often observed in cells, which requires further investigation to understand its cellular and physiological significance.trueATPases | Baculovirus | Chemomechanical | Kinesin-3 | Microtubule bending | SuperprocessiveKinesin-3 motors are fine-tuned at the molecular level to endow distinct mechanical outputsArticlehttps://bmcbiol.biomedcentral.com/counter/pdf/10.1186/s12915-022-01370-817417007December 20225177arJournal8