THE STRUCTURE OF THE SARCOMERIC M-BAND - LOCALIZATION OF DEFINED DOMAINS OF MYOMESIN, M-PROTEIN, AND THE 250-KD CARBOXY-TERMINAL REGION OF TITIN BY IMMUNOELECTRON MICROSCOPY

The M band of vertebrate cross-striated myofibrils has remained an eni gmatic structure. In addition to myosin thick filaments, two major str uctural proteins, myomesin and M-protein, have been localized to the M band. Also, titin is expected to be anchored in this structure. To be gin to understand the molecular layout of these three proteins, a pane l of 16 polyclonal and monoclonal antibodies directed against unique e pitopes of defined sequence was assembled, and immunoelectron microsco py was used to locate the position of the epitopes at the sarcomere le vel. The results allow the localization and orientation of defined dom ains of titin, myomesin, and M-protein at high resolution. The 250-kD carboxy-terminal region of titin clearly enters the M band with the ki nase domain situated similar to 52 nm from the central Mi-line. The po sitions of three additional epitopes are compatible with the view that the titin molecule reaches similar to 60 nm into the opposite sarcome re half. Myomesin also seems to bridge the central MI-line and is orie nted parallel to the long axis of the myofibril. The neighboring molec ules are oriented in an antiparallel and staggered fashion. The amino- terminal portion of the protein, known to contain a myosin binding sit e, seems to adopt a specific three-dimensional arrangement. While myom esin is present in both slow and fast fibers, M-protein is restricted to fast fibers. It appears to be organized in a fundamentally differen t manner: the central portion of the polypeptide is around the MI-line , while the terminal epitopes seem to be arranged along thick filament s. This orientation fits the conspicuously stronger M1-lines in fast t witch fibers. Obvious implications of this model are discussed.