Visualization of cardiac ventricular myosin heavy chain homodimers and heterodimers by monoclonal antibody epitope mapping

Two mAbs, one specific for cardiac alpha-myosin heavy chains (MHC) and the Other specific for cardiac beta-MHC, were used to investigate the heavy-chain dimeric organization of rat cardiac ventricular Myosin. Epitopes of the two mAbs were mapped on the Myosin molecule by electron microscopy of rotary shadowed mAb-myosin complexes. mAbs were clearly identifiable by the different locations of their binding sites on the Myosin rod. Thus, Myosin molecules could be directly discriminated according to their alpha-or beta-MHC content. alpha alpha-MHC and beta beta-MHC homodimers were visualized in complexes consisting of two molecules of the same mAb bound to one Myosin molecule. By simultaneously using the alpha-MHC-specific mAb and the beta-MHC-specific mAb, alpha beta-MHC heterodimers were visualized in complexes formed by one molecule of each of the two mAbs bound to one Myosin molecule. Proportions of alpha alpha-and beta beta-MHC homodimers and alpha beta-MHC heterodimers were estimated from quantifications of mAb-myosin complexes and compared with the proportions given by electrophoreses under nondenaturing conditions. This visualization of cardiac Myosin molecules clearly demonstrates the arrangement of alpha- and beta-MHC in alpha alpha-MHC homodimers, beta beta-MHC homodimers, and alpha beta-MHC heterodimers, as initially proposed by Hoh, J. F. Y., G. P. S. Yeoh, M. A. W. Thomas, and L. Higginbottom (1979).