title: Structure of the myosin filaments of relaxed and rigor vertebrate striated muscle studied by rapid freezing electron microscopy.
abstract: Rapid freezing followed by freeze-substitution has been used to study the ultrastructure of the myosin filaments of live and demembranated frog sartorius muscle in the states of relaxation and rigor. Electron microscopy of longitudinal sections of relaxed specimens showed greatly improved preservation of thick filament ultrastructure compared with conventional fixation. This was revealed by the appearance of a clear helical arrangement of myosin crossbridges along the filament surface and by a series of layer line reflections in computed Fourier transforms of sections, corresponding to the layer lines indexing on a 43 nm repeat in X-ray diffraction patterns of whole, living muscles. Filtered images of single myosin filaments were similar to those of negatively stained, isolated vertebrate filaments and consistent with a three-start helix. M-line and other non-myosin proteins were also very well preserved. Rigor specimens showed, in the region of overlapping myosin and actin filaments, periodicities corresponding to the 36, 24, 14.4 and 5.9 nm repeats detected in X-ray patterns of whole muscle in rigor; in the H-zone they showed a disordered array of crossbridges. Transverse sections, whose Fourier transforms extend to the (3, 0) reflection, supported the view, based on X-ray diffraction and conventional electron microscopy, that in the overlap zone of relaxed muscle most of the crossbridges are detached from the thin filaments while in rigor they are attached. We conclude that the rapid freezing technique preserves the molecular structure of the myofilaments closer to the in vivo state (as monitored by X-ray diffraction) than does normal fixation.
authors.affiliation: Department of Cell Biology University of Massachusetts Medical School, Worcester 01655.
authors.forename: R
authors.initials: R
authors.lastname: Craig
authors.forename: L
authors.initials: L
authors.lastname: Alamo
authors.forename: R
authors.initials: R
authors.lastname: Padrón
mesh_terms: D000818:Animals; D017110:Freeze Substitution; D047348:Hydrolyzable Tannins; D008854:Microscopy, Electron; D009119:Muscle Contraction; D009126:Muscle Relaxation; D009132:Muscles / Q000648:ultrastructure*; D009218:Myosins / Q000648:ultrastructure*; D011894:Rana pipiens; D011898:Ranidae; D014961:X-Ray Diffraction
chemical_list: D047348:Hydrolyzable Tannins; D009218:Myosins