Polymerization of actin: Mechanism of the Mg²⁺-induced process at pH 8 and 20°C

A detailed mechanism that fully accounts for the Mg²⁺-induced polymerization of actin in the presence or absence of Ca²⁺ at 20°C and pH 8 is presented. In the absence of Ca²⁺, the mechanism of the Mg²⁺-induced polymerization is as follows: Mg²⁺ binds to a metal-binding site on G-actin and induces a conformational change, which is required for eventual polymerization. The overall dissociation constant for this binding is about 30 μM. This actin species then binds a second molecule of Mg²⁺ (K(d) = 5 mM), which yields a species capable of polymerization. Dimer formation from this monomeric species is quite unfavorable, but trimer formation from dimer and monomer is much more favorable. The trimer may then elongate to give filaments. Ca²⁺ when present, binds at the same site as the tightly bound Mg²⁺ and must be displaced by Mg²⁺ before the conformational change can occur. The rate and dissociation constants for tight binding of Ca²⁺ and Mg²⁺ and for the conformational change are consistent with those observed previously by using a fluorescently labeled G-actin. With the mechanism proposed, it is possible to fit the full time course of polymerization over a wide range of actin concentrations, Mg²⁺ concentrations, and Ca²⁺ concentrations.