Estimation of Local Forces in Myofilaments using X-Ray Diffraction Patterns and Muscle Mechanics Data

Abstract

Small-angle X-ray diffraction of contracting muscle provide a view of sarcomere structure averaged over the area illuminated by the X-ray beam. Many important details of muscle contraction, however, such as crossbridge attachment and detachment rates are likely to depend on the local structures and forces on these structures. We have developed a methodology for predicting X-ray diffraction patterns with stepwise increases of strain along actin filaments in the hexagonal sarcomere lattice. Using PDB data for the crystal structures of G-actin we reconstructed the geometry of actin filaments deformed under stress generated by crossbridges. This stress varies along the filament length, so these deformations also vary with length. These fiber deformations may be determined by Monte Carlo calculations using the computational platform, MUSICO, along with the force and length changes seen during classical mechanical protocols. Using the predicted changes in spacings of actin filaments due to crossbridge forces we predicted how the X-ray diffraction patterns vary along with force and length during mechanical transients. Predicted X-ray diffraction patterns of fully developed isometric force muscle fibers are compared to experimental X-ray diffraction measurements in order to estimate the average force in actin filaments. Using peak shape analysis of the meridional reflections, one can assess the variation of filament deformation, and hence force, along the length of the filaments. Using the same methodology, we estimated the local modulation of force on actin during different mechanical protocols, including Huxley-Simmons quick releases and isotonic shortening. These data enable us to better correlate the molecular events, such as the current number of attached crossbridges and the distributions of crossbridge forces to macroscopic measurements of force and length changes. Supported by: NIH 9 P41 GM103622, R01 AR048776 and R01 DC 011528