Biology subjects

Shuman, H.

Publications and source records attributed to Shuman, H..

3 recordsLinked to original sources

bioRxiv2025

High resolution structures of Myosin-IC reveal a unique actin-binding orientation, ADP release pathway, and power stroke trajectory

Myosin-IC (myo1c) is a class-I myosin that supports transport and remodeling of the plasma membrane and membrane-bound vesicles. Like other members of the myosin family, its biochemical kinetics are altered in response to changes in mechanical loads that resist the power stroke. However, myo1c is unique in that the primary force-sensitive kinetic transition is the isomerization that follows ATP binding, not ADP release as in other slow myosins. Myo1c also powers actin gliding along curved paths, propelling actin filaments in leftward circles. To understand the origins of this unique force-sensing and motile behavior, we solved actin-bound myo1c cryo-EM structures in the presence and absence of ADP. Our structures reveal that in contrast with other myosins, the myo1c lever arm swing is skewed, partly due to a different actin interface that reorients the motor domain on actin. The structures also reveal unique nucleotide-dependent behavior of both the nucleotide pocket as well as an element called the N-terminal extension. We incorporate these observations into a model that explains why force primarily regulates ATP binding in myo1c, rather than ADP release as in other myosins. Integrating our cryo-EM data with available crystallography structures allows the modeling of full-length myo1c during force generation, supplying insights into its role in membrane remodeling. These results highlight how relatively minor sequence differences in members of the myosin superfamily can significantly alter power stroke geometry and force-sensing properties, with important implications for biological function. Significance StatementMyosin-IC (myo1c) uses an ATP-driven power-stroke to support slow intracellular membrane and vesicle transport. We used cryo-electron microscopy to understand adaptations of myo1c to perform its unique roles. We discovered an altered interface between myo1c and actin compared with the closely related myo1b, which repositions the motor domain and alters the trajectory of its lever arm swing compared to other myosins. This explains why myo1c propels actin filaments in a leftward circular path. We also discovered a unique role in force sensing for a structural element called the N-terminal extension and built a full-length atomic model for the myo1c power-stroke. Our findings highlight how myosins can tune their power-stroke geometries and force-sensing properties to adapt to diverse cellular functions.

biophysics↗

bioRxiv2020

Human hypertrophic cardiomyopathy mutation R712L suppresses the working stroke of cardiac myosin and can be rescued by omecamtiv mecarbil

Hypertrophic cardiomyopathies (HCMs) are the leading cause of acute cardiac failure in young individuals. Over 300 mutations throughout {beta}-cardiac myosin, including in the motor domain, are associated with HCM. A {beta}-cardiac myosin motor mutation (R712L) leads to a severe form of HCM. Actin-gliding motility of R712L-myosin is inhibited, despite near normal ATPase kinetics. By optical trapping, the working stroke of R712L-myosin was decreased 4-fold, but actin-attachment durations were normal. A prevalent hypothesis that HCM mutants are hypercontractile is thus not universal. R712 is adjacent to the binding site of the heart failure drug omecamtiv mecarbil (OM). OM suppresses the working stroke of normal {beta}-cardiac myosin, but remarkably, OM rescues the R712L-myosin working stroke. Using a flow chamber to interrogate a single molecule during buffer exchange, we found OM rescue to be reversible. Thus, the R712L mutation uncouples lever arm rotation from ATPase activity and this inhibition is rescued by OM.

biophysics↗

bioRxiv2019

Modulation of kinesin’s load-bearing capacity by force geometry and the microtubule track

Kinesin motors and their associated microtubule tracks are essential for long-distance transport of cellular cargos. Intracellular activity and proper recruitment of kinesins is regulated by biochemical signaling, cargo adaptors, microtubule associated proteins and mechanical forces. In this study, we found that the effect of opposing forces on the kinesin-microtubule attachment duration depends strongly on experimental assay geometry. Using optical tweezers and the conventional single-bead assay we show that detachment of kinesin from the microtubule is likely accelerated by forces vertical to the long-axis of the microtubule due to contact of the single bead with the underlying surface. We used the three-bead assay to minimize the vertical force component and found that when the opposing forces are mainly parallel to the microtubule the median attachment duration between kinesin and microtubules can be up to 10-fold longer than observed using the single-bead assay. Using the three-bead assay, we also found that not all microtubule protofilaments are equivalent interacting substrates for kinesin and that the median attachment duration (median-{Delta}t) of kinesin varies by more than 10-fold, depending on the relative angular position of the forces along the circumference of the microtubule. Thus, depending on the geometry of forces across the microtubule, kinesin can switch from a fast detaching motor (median-{Delta}t < 0.2 s) to a persistent motor that sustains attachment (median-{Delta}t > 3 s) at high forces (5 pN). Our data show that the load-bearing capacity of the kinesin motor is highly variable and can be dramatically affected by off-axis forces and forces across the microtubule lattice which has implications for a range of cellular activities including cell division and organelle transport.\n\nSignificance StatementKinesins are cytoskeletal motors responsible for the transport of cargoes along microtubules. It is well known that opposing forces decrease kinesins speed and run length. In this study, we found that when the pair of opposing forces applied on the kinesin-microtubule complex are parallel to the microtubule, the ability of kinesin to remain attached to the microtubule can vary by more than an order of magnitude depending on the relative azimuthal position of the pair of forces along the periphery of the microtubule. These results reveal a previously unknown versatility of kinesins load bearing capacity and as such have implications for the potential physiological roles of kinesin in a wide range of cell activities, including organelle transport and cell division.

biophysics↗