The optimized quantum dot mediated thermometry reveals isoform specific differences in efficiency of myosin extracted from muscle mini bundles

The biophysical function of myosin in vitro has been extensively investigated in different motility assays, but the study of myosin ATPase properties at the fiber level is insufficiently investigated. In this study, quantum dot (QD) mediated thermometry measurements were optimized to measure the efficiency of myosin extracted from muscle mini bundles. A reduction in fluorescent intensity of QD reflects an increase in temperature caused by the heat released during ATP hydrolysis and denotes the efficiency of the motor protein myosin. The procedure for extracting myosin was similar to the single fiber in vitro motility assay with some small modifications, and the concentration of myosin was represented by the extracted total protein since the ratio of extracted myosin to total protein was constant. Moreover, the efficiencies of myosin extracted from preparations containing different myosin heavy chain isoforms reveal lower efficiency of slow compared to fast myosin isoforms. Specifically, more heat was released in slow myosin enzymatic reaction, resulting in faster decay of QD fluorescence intensity. Hence, the optimized QD mediated thermometry provides a novel and sensitive approach to evaluate efficiency of myosin ATPase obtained from small muscle samples, representing a significant advantage in the clinical evaluation of neuromuscular disorders. The quantum dot (QD) thermometry has been optimized to measure myosin efficiency in minute amounts of muscle tissue. Myosin is prepared by the direct extraction from the muscle mini bundles rather than from pre-purified myosin. During ATP hydrolysis by myosin ATPase, heat is released, and temperature increases leading to a decreasing QD fluorescence intensity, indicating myosin efficiency. An isoform specific difference in myosin efficiency was observed with slow myosin having lower efficiency than fast myosin, as QD fluorescent intensity decayed more rapidly in slow myosin. [Display omitted] •Quantum dot mediated thermometry was optimized to evaluate the efficiency of myosin in vitro.•Myosin was extracted from muscle mini bundles in short time rather than from time and tissue consuming purification.•QD thermometry detected myosin isoform specific differences, as slow myosin demonstrates less efficiency.