Relation Between Crossbridge Structure and Actomyosin ATPase Activity in Rat Heart

From the Department of Physiology, School of Medicine, University of Pennsylvania, Philadelphia.

Correspondence to Saul Winegrad, MD, Department of Physiology, School of Medicine, University of Pennsylvania, 37th and Hamilton Walk, Philadelphia, PA 19104-6085. E-mail bsg{at}mail.med.upenn.edu

Abstract—Cardiac myofilaments contain proteins that regulate the interaction between actin and myosin. In the thick filament, there are several proteins that may contribute to the regulation of the contraction. The myosin binding protein C, or C protein, has 4 sites that can be phosphorylated by a Ca²⁺-calmodulin–controlled kinase, protein kinase A or protein kinase C. Using electron microscopy and optical diffraction, we examined the structure of thick filaments isolated from rat ventricles with either the or ß isoform of myosin heavy chain (MHC) and the effect of specific phosphorylation of C protein on the structure. In thick filaments with -MHC, crossbridges were clearly visible. Phosphorylation of C protein by protein kinase A extended the crossbridges from the backbone of the filament, changed their orientation, increased the degree of order of the crossbridges, and decreased the flexibility of the crossbridges. Crossbridges in filaments with ß-MHC were less ordered and apparently more flexible. Phosphorylation of C protein in ß-MHC–containing filaments did not extend the crossbridges and did not alter degree of order or flexibility. The relative flexibility of the crossbridges inferred from the optical diffraction pattern correlated well with the rate of ATP hydrolysis by actomyosin. These results suggest that (1) crossbridge flexibility is an important parameter in setting the rate of crossbridge cycling, and (2) C protein–mediated control of the position and flexibility of crossbridges may regulate actomyosin ATPase activity by modifying the kinetics of crossbridge cycling.

Key Words: crossbridge • ATPase • C protein • myosin • phosphorylation

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