Abstract
We have attached a spin label or a fluorescent dye to actin to probe the interaction of G-actin with myosin subfragments. The spin label spectra, which are sensitive to the polymerization of the actin, showed that when heavy meromyosin was added to G-actin the actin was polymerized. When actin was in excess, the spin label spectra indicated that 3.8 ± 0.6 actin monomers were polymerized per heavy meromyosin molecule. Addition of myosin subfragment-1 on the other hand caused a negligible amount of actin polymerization as measured by the spectra of the spin label attached to the actin. The polarization of the fluorescence of the dye bound to actin increased upon addition of heavy meromyosin to the G-actin. In good agreement with the spin label results, the fluorescence data indicated that, when actin was in excess, 4.2 actin monomers were incorporated into a polymer per heavy meromyosin molecule added. One heavy meromyosin per two actin monomers was necessary and sufficient to “polymerize” all of the actin. When heavy meromyosin was added to succinylated actin, which was incapable of polymerization, the same changes in the fluorescent polarization were observed as with unsuccinylated actin. Addition of subfragment-1 to fluorescent-labeled actin indicated that subfragment-1 formed a 1:1 molar complex with the actin monomers. These data are interpreted in terms of a model in which the heavy meromyosin and actin form a copolymer which involves cooperative interactions between the two heads of heavy meromyosin.
Published Version
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