A mixed-kinetic model describes unloaded velocities of smooth, skeletal, and cardiac muscle myosin filaments in vitro.

In vitro motility assays, where purified myosin and actin move relative to one another, are used to better understand the mechanochemistry of the actomyosin adenosine triphosphatase (ATPase) cycle. We examined the relationship between the relative velocity () of actin and myosin and the number of available myosin heads () or [ATP] for smooth (SMM), skeletal (SKM), and cardiac (CMM) muscle myosin filaments moving over actin as well as from actin filaments moving over a bed of monomeric SKM. These data do not fit well to a widely accepted model that predicts that is limited by myosin detachment from actin (/), where equals step size and equals time a myosin head remains attached to actin.

Myosin light chain kinase steady-state kinetics: comparison of smooth muscle myosin II and nonmuscle myosin IIB as substrates.

Unlabelled: Myosin light chain kinase (MLCK) phosphorylates S19 of the myosin regulatory light chain (RLC), which is required to activate myosin's ATPase activity and contraction. Smooth muscles are known to display plasticity in response to factors such as inflammation, developmental stage, or stress, which lead to differential expression of nonmuscle and smooth muscle isoforms. Here, we compare steady-state kinetics parameters for phosphorylation of different MLCK substrates: (1) nonmuscle RLC, (2) smooth muscle RLC, and heavy meromyosin subfragments of (3) nonmuscle myosin IIB, and (4) smooth muscle myosin II.

Diffusion of myosin light chain kinase on actin: A mechanism to enhance myosin phosphorylation rates in smooth muscle.

Smooth muscle myosin (SMM) light chain kinase (MLCK) phosphorylates SMM, thereby activating the ATPase activity required for muscle contraction. The abundance of active MLCK, which is tightly associated with the contractile apparatus, is low relative to that of SMM. SMM phosphorylation is rapid despite the low ratio of MLCK to SMM, raising the question of how one MLCK rapidly phosphorylates many SMM molecules.

Velocities of unloaded muscle filaments are not limited by drag forces imposed by myosin cross-bridges.

It is not known which kinetic step in the acto-myosin ATPase cycle limits contraction speed in unloaded muscles (V0). Huxley's 1957 model [Huxley AF (1957) Prog Biophys Biophys Chem 7:255-318] predicts that V0 is limited by the rate that myosin detaches from actin. However, this does not explain why, as observed by Bárány [Bárány M (1967) J Gen Physiol 50(6, Suppl):197-218], V0 is linearly correlated with the maximal actin-activated ATPase rate (vmax), which is limited by the rate that myosin attaches strongly to actin.