Harmonic Force Spectroscopy Reveals a Force-Velocity Curve from a Single Human Beta Cardiac Myosin Motor

A muscle contracts rapidly under low load, but slowly under high load. This
load-dependent muscle shortening has been described with a hyperbolic
load-velocity curve. Its molecular mechanisms remain to be elucidated, however.
During muscle contraction, myosins in thick filaments interact with actin in
thin filaments in the sarcomere, cycling between a strongly bound state (force
producing state) and a weakly bound state (relaxed state). Huxley and Simmons
have previously proposed that the transition from the strong to the weak interaction can be modulated by an external load, i.e., the transition is slow under
high load and fast under low load.
We use a new, simple method we call "harmonic force spectroscopy" to
extract a load-velocity relationship from a single human beta cardiac myosin
II motor (S1). With a dual-beam optical trap, we hold an actin dumbbell over a
single myosin molecule that is anchored to the microscope stage, which we
oscillate sinusoidally in the direction of the dumbbell. Upon binding of the
motor to the actin filament, it experiences an oscillatory load with a mean
value that may be directed forward or backward, depending on where the binding took place. We find that the duration of the strongly bound state at saturating [ATP] is exponentially correlated with the mean load applied to the
myosin during the bound state, which is explained by Arrhenius transition theory.
With an independent stroke size measurement, we obtained an exponential
load-velocity curve from a single human beta cardiac myosin S1. We also
compare load-velocity curves for wild-type motors with load-velocity curves
of mutant forms that cause hypertrophic or dilated-cardiomyopathy (HCM
or DCM), in order to understand the effects of mutations on the contractile cycle
at the single molecule level.