What Determines the Directionality of Kinesins? Studying the Anomalous Directionality of Nonclaret Disjunctional Motor Mutants

NCD (nonclaret disjunctional) motors are a class of kinesin
family
proteins that are known to walk toward the minus end along the microtubule.
Understanding the origin of such unique directionality of NCD motors
and related kinesins has been a problem of fundamental importance.
Experimental studies have indicated that specific NCD motor mutants
switch their directionality. A type of NCD mutant (ran12) walks in
the opposite direction (plus end) compared to the wild type (WT),
whereas another mutant (N340K) of NCD shows diffusive bidirectionality.
This work establishes that the directionality of these NCD motors
is regulated by the barrier of the rate-determining ADP release step.
We explore the molecular origin of the anomalous directionality of
the mutated NCD motors compared with the wild type and determine the
change in the barriers of ADP release for forward and backward motions.
Our investigation is performed by combining different simulation techniques
such as coarse-grained (CG) modeling, all-atom steered molecular dynamics
(SMD) simulation, free energy estimation, as well as structural and
stability analysis, and the maximum entropy (MaxEnt) method. We conclude
that the differences in the directionality of different types of NCD
motors are controlled by the free energy barrier for the ADP release
step. Finally, we show that the statistical energy deduced from homologous
sequences by the maximum entropy (MaxEnt) principle is significantly
correlated with the stability and experimentally measured velocities
of the wild type and several NCD mutants, and established that the
stability loss of specific NCD mutants contributes to their reduced
velocities and anomalous directionality on the microtubules. This
provides a rare case where the origin of the AI prediction is clearly
related to a physical-based picture.