The kinetics and mechanism of myosin subfragment-1-induced polymerization of G-actin into F-actin-S1-decorated filaments have been investigated in low ionic strength buffer and in the absence of free ATP. The mechanism of assembly of F-actin-S1 differs from salt-induced assembly of F-actin. Initial condensation of G-actin and S1 into oligomers in reversible equilibrium is a prerequisite step in the formation of F-actin-S1 . Oligomers have a relatively low stability (10(6) M-1) and contain S1 in a molar ratio to actin close to 0.5. Increased binding of S1 up to a 1:1 molar ratio to actin is associated with further irreversible condensation of oligomers into large F-actin-S1 structures of very high stability. In contrast to salt-induced assembly of F-actin, no monomer-polymer equilibrium, characterized by a critical concentration, can be defined for F-actin-S1 assembly, and end-to-end annealing of oligomers is predominant over growth from nuclei in the kinetics. Simultaneous recordings of the changes in light scattering, pyrenyl- and NBD-actin fluorescence, ATP hydrolysis, and release of Pi during the polymerization process have been analyzed to propose a minimum kinetic scheme for assembly, within which several elementary steps, following oligomer formation, are required for assembly of F-actin-S1. ATP hydrolysis occurs before polymerization of MgATP-G-actin but not of CaATP-G-actin. The release of inorganic phosphate occurs on F-actin-S1 at the same rate as on F-actin.