We have recently discovered that myosin crossbridges bind to actin in relaxed muscle and interact in a manner characterized by rapid, reversible attachment and detachment. We have also found that the rate of crossbridge attachment and detachment depends upon the nucleotide or nucleotide analogue bound to the active site. The attachment/detachment rates are rapid in the presence of ATP, somewhat more moderate in the presence of pyrophosphate (PPi) or adenyl-5-yl imidodiphosphate (AMPPNP), and slow in the presence of ADP. We detect crossbridge detachment by measuring the decay of tension following rapid stretch of an isolated single muscle fiber which has had its outer membrane removed to facilitate equilibration with experimental solutions. Using thermodynamic relationships derived by T. L. Hill, we have been able to show that, whereas the crossbridge attachment rate-constants are difficult to derive, mechanical experiments can provide considerable information about the detachment rate-constants. We found the detachment rate-constants to be greater than 5x1000 s-1 in the presence of ATP, about 0.1-10 s-1 in the presence of PPi, and less than 0.001 s-1 in the presence of ADP. For a given nucleotide concentration, detachment occurs not with a single rate-constant but with a range of rate-constants. Recently we have been able to show that some of this range is related to strain-sensitivity of the rate-constants since not all attached cross-bridges are equally strained. These experiments show that, to a first approximation, crossbridge behavior can be understood in terms of a fairly simple attachment/detachment equilibrium. In addition, some of the relevant rate-constants pertinent to the crossbridge contractile cycle have been worked out.
