Elementary Steps of Contraction Probed by Sinusoidal Analysis Technique in Rabbit Psoas Fibers

Abstract

Elementary steps of contraction were probed by sinusoidal analysis technique in skinned fibers from the rabbit psoas muscle during maximal Ca2+ activation (pCa 4.55-4.82) at 20 degrees C and 200 mM ionic strength. Our study included the effects of MgATP, MgADP, and Pi concentrations, and an ATP hydrolysis rate measurement. We increased the frequency range up to 350 Hz, and resolved an extra process (D), in addition to well defined processes (A), (B), and (C). Based on these studies, we established a cross-bridge scheme consisting of six attached states, one detached state, and transitions between these states. We deduced all kinetic constants to specify the scheme. The scheme uniquely explains our data, and no other scheme with an equal degree of simplicity could explain our data. We correlated process (D) to ATP isomerization, process (C) to cross-bridge detachment, and process (B) to cross-bridge attachment. We deduced the tension per cross-bridge state, which indicates that force is generated on cross-bridge attachment and before Pi-release. We also found that the rate constants of elementary steps become progressively slower starting from ATP binding to the myosin head and ending by ADP isomerization, and this stepwise slowing may be the essential and integral part of the energy transduction mechanism by muscle.