Ionic Strength Influences the Mechanical Force Regulation of Myosin'S Unbinding Rate

Abstract

We previously showed that skeletal muscle actomyosin in various nucleotide states behaves as a catch bond and that bond lifetime is maximal at the isometric force generated by a single myosin molecule. Recent data from our lab suggest the catch-slip behaviour results from allosteric conversion of myosin from a short- to a long-lived bound state. Though these studies shed some light on actin-myosin bond mechanics, the experimental conditions were 5 times lower than physiologic ionic strength. We therefore investigated the possibility that residence times in the short- and long-lived bond state of the myosin head are influenced by ionic strength. Dynamic force spectroscopy was used to measure the load-dependent bond dissociation of nucleotide-free heavy meromyosin (HMM) from actin at 25 mM (typical in vitro) and 145 mM KCl (physiologic ionic strength). In this technique, a trapped actin-coated bead was brought into contact with an HMM-coated surface, a bond was allowed to form, and then the bond was loaded until rupture by moving the laser trap away. We also determined bond lifetimes at near-zero load by placing an actin-coated bead adjacent to the HMM-coated surface and allowing bonds to form and break semi-spontaneously. Actin-myosin bond lifetimes were higher on average at physiologic ionic strength compared to low ionic strength across a range of forces. Further, our data suggest that catch bond behaviour is abrogated at physiologic ionic strength.