Annotating the x-ray diffraction pattern of vertebrate striated muscle.

Low-angle x-ray diffraction is a powerful technique for analyzing the molecular structure of the myofilaments of striated muscle in situ. It has contributed greatly to our understanding of the relaxed, 430-Å repeating organization of myosin heads in thick filaments in skeletal and cardiac muscle. Using x-ray diffraction, changes in filament structure can be detected on the angstrom length scale and millisecond timescale, leading to models that are the foundation of our understanding of the structural basis of contraction. As with all x-ray fiber diffraction studies, interpretation requires modeling, which has previously been based on low-resolution knowledge of thick filament structure and is complicated by the contributions of multiple filament components to most x-ray reflections. Here, we use an atomic model of the human cardiac thick filament C-zone, derived from cryo-EM in the presence of the myosin inhibitor, mavacamten, to compute objectively the contributions of myosin heads, tails, titin, and cMyBP-C to the diffraction pattern, by including/excluding these components in the calculations. Our results support some previous interpretations but contradict others. We confirm that the myosin heads are responsible for most of the intensity on the myosin layer lines, including the M3 meridional. Contrary to expectation, we find that myosin tails contribute little to the pattern, including the M6 meridional; this reflection arises mainly from heads and other components. The M11 layer line (39-Å spacing) arises mostly from the curved and kinked structure of titin, which allows 11 ∼42-Å-long domains to fit into the 430-Å repeat. The M11 spacing can be used as a measure of strain in the myosin filament backbone as there is negligible head contribution. The computed layer lines account well for the experimentally determined pattern. These insights should aid future understanding of the x-ray pattern of intact muscle in different conditions such as contraction and drug treatment.