AXIAL ROTATION OF SLIDING ACTIN-FILAMENTS REVEALED BY SINGLE-FLUOROPHORE IMAGING

In the actomyosin motor, myosin slides along an actin filament that ha s a helical structure with a pitch of approximate to 72 nm, Whether my osin precisely follows this helical track is an unanswered question be aring directly on the motor mechanism, Here, axial rotation of actin f ilaments sliding over myosin molecules fixed on a glass surface was vi sualized through fluorescence polarization imaging of individual tetra methylrhodamine fluorophores sparsely bound to the filaments. The fila ments underwent one revolution per sliding distance of approximate to 1 mu m, which is much greater than the 72 nm pitch, Thus, myosin does not ''walk'' on the helical array of actin protomers; rather it ''runs ,'' skipping many protomers, Possible mechanisms involving sequential interaction of myosin with successive actin protomers are ruled out at least for the preparation described here in which the actin filaments ran rather slowly compared with other in vitro systems, The result al so indicates that each ''kick'' of myosin is primarily along the axis of the actin filament, The successful, real-time observation of the ch anges in the orientation of a single fluorophore opens the possibility of detecting a conformational change(s) of a single protein molecule at the moment it functions.