Identification of functional differences between recombinant human α and β cardiac myosin motors

The myosin isoform composition of the heart is dynamic in health and disease and has been shown to affect contractile velocity and force generation. While different mammalian species express different proportions of alpha and beta myosin heavy chain, healthy human heart ventricles express these isoforms in a ratio of about 1:9 (alpha:beta) while failing human ventricles express no detectable alpha-myosin. We report here fast-kinetic analysis of recombinant human alpha and beta myosin heavy chain motor domains. This represents the first such analysis of any human muscle myosin motor and the first of alpha-myosin from any species. Our findings reveal substantial isoform differences in individual kinetic parameters, overall contractile character, and predicted cycle times. For these parameters, alpha-subfragment 1 (S1) is far more similar to adult fast skeletal muscle myosin isoforms than to the slow beta isoform despite 91% sequence identity between the motor domains of alpha- and beta-myosin. Among the features that differentiate alpha- from beta-S1: the ATP hydrolysis step of alpha-S1 is similar to ten-fold faster than beta-S1, alpha-S1 exhibits similar to five-fold weaker actin affinity than beta-S1, and actin center dot alpha-S1 exhibits rapid ADP release, which is > ten-fold faster than ADP release for beta-S1. Overall, the cycle times are ten-fold faster for alpha-S1 but the portion of time each myosin spends tightly bound to actin (the duty ratio) is similar. Sequence analysis points to regions that might underlie the basis for this finding.