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Identification of functional differences between recombinant human α and β cardiac myosin motors
John C Deacon1, Marieke J Bloemink, Heresh Rezavandi
1Department of Molecular, Cellular and Developmental Biology and Biofrontiers Institute, University of Colorado, MCDB, Boulder, CO 80309, USA.
Insights
Human heart myosin isoforms, alpha and beta, show significant kinetic differences. Alpha-myosin motor domains function similarly to fast skeletal muscle, despite high sequence identity with beta-myosin.
Area of Science:
- Biochemistry
- Molecular Biology
- Cardiology
Background:
- Myosin isoform composition in the heart is crucial for contractile function and changes in disease states.
- Human ventricles typically express a 1:9 ratio of alpha- to beta-myosin heavy chain, with alpha-myosin absent in heart failure.
Purpose of the Study:
- To perform fast-kinetic analysis on recombinant human alpha- and beta-myosin heavy chain motor domains.
- To compare the kinetic properties and contractile characteristics of human alpha- and beta-myosin motor domains.
Main Methods:
- Fast-kinetic analysis of recombinant human alpha- and beta-myosin heavy chain motor domains (subfragment 1).
- Measurement of ATP hydrolysis rates, actin binding affinity, and ADP release rates.
Main Results:
- Alpha-myosin subfragment 1 (S1) demonstrated a ~ten-fold faster ATP hydrolysis step compared to beta-S1.
- Alpha-S1 exhibited ~five-fold weaker actin affinity and significantly faster ADP release than beta-S1.
- Despite kinetic differences, the duty ratio (time bound to actin) was similar for both isoforms, with alpha-S1 having ten-fold faster overall cycle times.
Conclusions:
- Human alpha- and beta-myosin motor domains possess distinct kinetic properties influencing cardiac contractility.
- Alpha-myosin's kinetic profile resembles fast skeletal muscle myosin, suggesting functional specialization.
- Identified kinetic differences may be attributed to specific sequence variations within the myosin motor domains.
Abstract:
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 α and β myosin heavy chain, healthy human heart ventricles express these isoforms in a ratio of about 1:9 (α:β) while failing human ventricles express no detectable α-myosin. We report here fast-kinetic analysis of recombinant human α and β myosin heavy chain motor domains. This represents the first such analysis of any human muscle myosin motor and the first of α-myosin from any species. Our findings reveal substantial isoform differences in individual kinetic parameters, overall contractile character, and predicted cycle times. For these parameters, α-subfragment 1 (S1) is far more similar to adult fast skeletal muscle myosin isoforms than to the slow β isoform despite 91% sequence identity between the motor domains of α- and β-myosin. Among the features that differentiate α- from β-S1: the ATP hydrolysis step of α-S1 is ~ten-fold faster than β-S1, α-S1 exhibits ~five-fold weaker actin affinity than β-S1, and actin·α-S1 exhibits rapid ADP release, which is >ten-fold faster than ADP release for β-S1. Overall, the cycle times are ten-fold faster for α-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.
