Biophysical Society Thematic Meeting | Canterbury 2023

Towards a More Perfect Union: Multi-Scale Models of Muscle and Their Experimental Validation

Monday Speaker Abstracts

EVIDENCE FOR TWO DISTINCT ACTOMYOSIN CROSS-BRIDGE STIFFNESS FOR HUMAN BETA CARDIAC MYOSIN Tianbang Wang 1 ; Theresia Kraft 1 ; Arnab Nayak 1 ; Mamta Amrute-Nayak ; 1 Medical School Hannover, Molecular and Cell Physiology, Hannover, Germany Stiffness, i.e., resistance to elastic deformation, is a crucial physical parameter that determines the force-generation and force-sensing ability of the motor proteins, such as cardiac myosin II. During muscle or cardiac contraction, an elastic element within the actomyosin (AM) cross bridge allows the strain to develop, and the release of this strain then drives the actin filament sliding relative to the myosin or thick filament. Here, we employed a single-molecule optical trapping method to analyze the rigidity of native human ventricular b-cardiac myosin (b-CM). The intermittent interactions between the actin and myosin in the presence of ATP were analyzed using the variance-covariance method (Lewalle et al., 2008), to determine the stiffness of the myosin. Interestingly, we found that the measured stiffness of the b-CM was sensitive to the ATP concentration. With increasing [ATP] the AM cross-bridge stiffness decreased, suggesting that the stiffness was dependent on the nucleotide state as the myosin progressed through the ATPase cycle. The strongly bound AM.ADP state exhibited > two-fold lower stiffness (~0.6 pN/nm) than the AM rigor state (~1.9 pN/nm). We further confirm the high stiffness value as rigor stiffness by employing pyrophosphate (PPi) in our assays. In the presence of PPi, the actin and myosin underwent binding and unbinding events. The bound state, i.e., nucleotide-free state, thus represented the near-rigor AM state with the stiffness of ~ 2 pN/nm. We speculate that more compliant AM.ADP state may be important for the crossbridge response to assisting or resisting load in cardiac myosins and other myosin forms where ADP release is known to be load sensitive. Our studies revealed previously unexplored aspects of myosin’s mechanical property. This information is vital to understand cardiac disorders such as hypertrophic cardiomyopathy linked to point mutations in b-CM components that alter the rigidity of ventricular myosin.

MYOSIN-BINDING PROTEIN C/H: IMPACT ON MUSCLE CONTRACTILITY AND DEVELOPMENT David Warshaw University of Vermont, USA No Abstract

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