Biophysical Society Thematic Meeting | Canterbury 2023

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

Poster Abstracts

23-POS Board 23 HCM- CAUSING MUTATIONS G256E AND G768R IN Β -CARDIAC MYOSIN CAUSE HYPERCONTRACTILITY BY OPENING MYOSIN HEADS Divya Pathak 1,2 ; Aminah Dawood 1 ; Kathleen Ruppel 1,2 ; James A Spudich 1,2 ; Hypertrophic cardiomyopathy (HCM) is an inherited cardiovascular disease that affects up to 1 in 200 people and is the leading cause of sudden cardiac death among the young. HCM is marked by hypercontractility, abnormal thickening of the heart muscle, cardiac hypertrophy and fibrosis. Over 30% of all known HCM mutations can be found in the sarcom ere protein β -cardiac myosin, but their molecular mechanism remains poorly understood. β -cardiac myosin is a large, highly allosteric protein, making it challenging to predict functional changes due to mutations. Studies over the past decade have shown that myosin samples two conformational states– an ‘open’ state, where the myosin heads are available to interact with actin and a ‘closed’ state, where the heads fold back onto the myosin tail and are no longer available to interact with actin. We have previously shown that several HCM mutations cause hypercontractility by increasing the myosin heads in the ‘open’ state. This study focuses on two point mutations in distinct regions of the myosin motor domain – G256E and G768R. G256 is located in the hairpin turn in the myosin transducer region bordering the myosin mesa, whereas G768 is present at the end of its converter in the motor domain of β -cardiac myosin. Both mutations have been predicted to destabilize the ‘closed’ or folded-back state. I will be presenting the characterization of these mutations using single-molecule and ensemble assays. Although the velocity for G256E decreased by ~20%, the mutation increased the number of myosin in ‘open’ state, resulting in hypercontractility. Interestingly, G768R led to a steep decrease of ~80% in actin velocity in an in vitro actin gliding assay. On a single molecule level, this mutation significantly decreased myosin’s detachment rate without altering force sensitivity. Despite decreasing the velocity and ATPase activity per motor head, I will present results showing how the mutation compensates for the reduced ATPase activity by biasing the myosin equilibrium towards the ‘open state’. 1 Stanford University, Biochemistry, Stanford, CA, USA 2 Stanford Cardiovascular Institute, Stanford, CA, USA

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