Kinetic analysis of actin polymerization.

The kinetics of actin polymerization were analyzed by measuring the changes in absorbance which accompany the G-F transformation of actin. In these studies, gel-filtered actin was polymerized in the absence of shearing stress under physiological ionic conditions. Self-polymerization was found to be characterized as a process having a lag phase followed by a pseudo-first-order decay process, suggesting that actin polymerization consists of distinct nucleation and elongation phases. The size of the nucleus was estimated to be two to four monomer units by analyzing the actin concentration-dependences of the rate constant for the pseudo-first-order process, the maximal rate of polymerization and the half-polymerization time. The elongation reaction was induced by mixing actin filament seeds with actin monomers under conditions where spontaneous nucleation is slow. This elongation was also found to be a pseudo-first-order process, indicating that spontaneous nucleation was negligible. Our data suggested that the initial rate of elongation was proportional to both the number concentration of actin filaments and the actin monomer concentration above the critical concentration, and that the depolymerization rate was proportional to the number concentration of actin filaments but independent of monomer concentration. The results of direct analysis of the depolymerization reaction were consistent with this suggestion. These studies strongly support the condensation polymerization mechanism as a model for actin polymerization.