Position of Mercenaria regulatory light-chain Cys50 site on the surface of myosin visualized by electron microscopy

Mercenaria regulatory light-chains, specifically labelled at cysteine 50 with N-iodoacetyl-N'-biotinylhexylenediamine, were rebound to regulatory light-chain denuded scallop myosin, and the hybrid myosin formed was decorated with avidin. These hybrid myosins were visualized by rotary-shadowing electron microscopy. Three distinct images of avidin-decorated hybrid myosin molecules were obtained. These comprise singly decorated molecules, where the avidin is bound symmetrically or asymmetrically with respect to the two heads of myosin, in addition to "figures-of-five", where two myosin molecules associate with a centrally placed avidin molecule. Analysis of these images indicates that the Mercenaria regulatory light-chain Cys50 site is located 15 to 35 A from the head-rod junction when the light-chain is bound in situ to myosin. Implications with respect to head topology and probe studies are discussed.