Activation and Inhibition of F-Actin and Cardiac Thin Filaments by the N-Terminal Domains of Cardiac Myosin Binding Protein C

Abstract

Myosin binding protein-C (MyBPC) has been known for over thirty years to inhibit actomyosin ATP hydrolysis and more recently has been shown to affect the calcium sensitivity of force in muscle fibers. The various domains of MyBPC have previously been shown to have complex interactions with other myofibrillar proteins. The C-terminal domains have been shown to bind to the thick filament and the N-terminal domains interact with the S2 region of myosin and with f-actin. The number of mutations in cardiac MyBPC (cMyBPC) are the second most prevalent of sarcomeric proteins in producing cardiomyopathies. We have used steady state kinetics to study the molecular mechanism by which the soluble N-terminal C0C1, C1C2 and C0C2 domains of mouse and human cMyBPC affect the activation of myosin ATP hydrolysis by f-actin and native porcine thin filaments.The N-terminal domains of cMyBPC inhibit the activation of the steady state ATPase myosin by f-actin. However, in the presence of native cardiac thin filaments.the N-terminal domains C1C2 and C0C2 of mouse and human cMyBPC produce biphasic effects on the steady state ATP hydrolysis of cardiac myosin-S1. That is, ATPase is activated at low ratios of cMyBPC-N-terminal domain to thin filament and is inhibited by higher ratios similar to the effects observed with f-actin. These data suggest that low ratios of cMyBPC N-terminal domains activate thin filaments in a mechanism similar to that of rigor myosin-S1 but higher ratios inhibit the ATPase rate by competing with myosin-S1-ADP-Pi binding to actin and thin filaments. Effects also appear species-dependent since the C0C1 domains of human cMyBPC produce similar effects as C1C2 and C0C2 on ATPase in the presence of thin filaments, but mouse C0C1 does not produce significant activation.