Probing myosin head structure with monoclonal antibodies.
Monoclonal antibodies that react with defined regions of the heavy and light chains of chicken skeletal muscle myosin have been used to provide a correlation between the primary and the tertiary structures of the head. Electron microscopy of rotary shadowed antibody-myosin complexes shows that the sites for three epitopes in the 25,000 Mr tryptic fragment (25k) of subfragment-1, including one within 4000 Mr of the amino terminus of the myosin heavy chain, are clustered 145(+/- 20) A from the head-rod junction. An epitope in the 50,000 Mr fragment maps even further out on the head. These antibodies bind to the head in several orientations, suggesting that each of the heads can rotate can rotate 180 degrees about the head-rod junction. The epitopes are accessible on subfragment-1 bound to actin when they were probed with Fab fragments; therefore, none of these heavy chain sites is is on the contact surface between the head and actin. Two of the anti-25k antibodies affect the K+-EDTA-and Ca2+-ATPase activities of myosin in a manner that mimics the effect on activity of the modification of the reactive thiol, SH-1. These two antibodies also inhibit the actin-activated ATPase non-competitively with respect to actin. None of the other eight antibodies tested had any marked effect on activity. A monoclonal antibody that reacts with an epitope in the amino-terminal third of myosin light chain 2 maps close to the head-rod junction. A polyclonal antibody specific for the amino terminus of light chain 3 binds further up in the "neck region" of the head, indicating that these portions of the two classes of light chains are located at different sites.
D000251:Adenosine Triphosphatases / Q000378:metabolism; D000818:Animals; D000911:Antibodies, Monoclonal; D001666:Binding Sites, Antibody; D002645:Chickens; D007140:Immunoglobulin Fab Fragments / Q000276:immunology; D007074:Immunoglobulin G / Q000276:immunology; D008854:Microscopy, Electron; D009218:Myosins / Q000276:immunology; D011487:Protein Conformation
D000911:Antibodies, Monoclonal; D007140:Immunoglobulin Fab Fragments; D007074:Immunoglobulin G; D000251:Adenosine Triphosphatases; D009218:Myosins