Conformational Dynamics of Nucleotide-Free Myo1b

Myo1b is the widely expressed myosin-I isoform with kinetics that are highly sensitive to tension (Laakso et al. 2008. Science. 321:133-6). To better understand the structural transitions that accompany the myo1b power stroke, we analyzed the displacement of actin filaments by single myo1b molecules in a 3-bead trap assay. Under conditions where we predict myo1b to be nucleotide-free, we observe “flickering” of the position of myo1b-bound actin between two distinct states. We are able to measure the rate of transition between these states, and we have found the rates to be force sensitive. Interestingly, previous biochemical kinetic measurements have shown that actin-bound myo1b (in the absence of nucleotide) is in equilibrium between a state that does not bind nucleotide (AM) and a state can bind nucleotide (AM’). The rate of the transition from the AM to AM’ state is similar to the rate of ADP release, and it was proposed (Geeves et al. 2000. J. Biol. Chem. 275:21624-30) that this conformational transition is similar to the conformational change that accompanies ADP release. Our observed flickering rates are similar to the AM to AM’ transitions rates. Because we found ADP release to be the force-sensitive transition, it is interesting to consider the role of this nucleotide-free transition in myo1b tension-sensing.