The energetics of molecular motors

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Actin and myosin are two proteins that make up molecular motors that drive many functions in animals including cell division, ATP production, regulation of hearing, and most importantly contractions in striated (skeletal and heart) muscles. We have studied the molecular mechanisms of muscle contractions for the past 15 years and realized that the textbook version of muscle contraction cannot explain many of the experimentally observed phenomena. Specifically, when an active muscle is stretched, its force, energetics and stiffness are not explained by the molecular motor comprised of actin and myosin. Just over a decade ago, we found that structural proteins, thought to only play passive supportive roles, contribute to active force regulation and contraction. We identified through molecular, sub-cellular and cellular experiments that the structural element contributing to active force regulation is the molecular spring titin. We suggest, and have preliminary evidence, that titin binds calcium upon muscle activation, thereby increasing its inherent stiffness and force, and that titin binds to actin, thus shortening its free spring length, thereby increasing its stiffness and force upon stretch. If our theories are correct, then the energy cost (ATP, adenosine triphosphate) per unit of force should be much smaller during and after stretch of active muscle compared to the cost of the corresponding force following passive muscle stretching. However, the energetic costs of single myofibrils and single muscle cells have never been measured, thus our predictions cannot be tested experimentally at this time. The purpose of this application is to acquire an inverted fluorescent microscope and a single cell mechanical testing apparatus to be integrated into an existing single myofibril system that allows for simultaneous mechanical and energetic property measurements while the actin-myosin motors are activated through calcium and ATP. This system would allow for continuous measurement of the force, stiffness and ATP use in single myofibrils and cardiac and skeletal muscle cells. To our best knowledge, such a system does not exist anywhere and there are definitely no accounts of the energetics of single myofibril contraction published in the literature.