The effect of profilin, a G-actin binding protein, on the mechanism of exchange of the tightly bound metal ion and nucleotide on G-actin, has been investigated. 1) In low ionic strength buffer, profilin increases the rates of Ca and Mg dissociation from G-actin 250- and 50-fold, respectively. On the profilin-actin complex as well as on G-actin alone, nucleotide exchange is dependent on the concentration of divalent metal ion and is kinetically limited, at low concentration of metal ion, by the dissociation of the metal ion. 2) Under physiological ionic conditions, nucleotide exchange on G-actin is 1 order of magnitude faster than at low ionic strength. The rate of MgATP dissociation is increased by profilin from 0.05 s to 2 s, the rate of MgADP dissociation is increased from 0.2 s to 24 s. The dependences of the exchange rates on profilin concentration are consistent with a high affinity (5 10 to 10M) of profilin for ATP-G-actin, and a 20-fold lower affinity for ADP-Gactin. Profilin binding to actin lowers the affinity of metal-nucleotide by about 1 order of magnitude. These results restrain the possible roles of profilin in actin assembly in vivo.

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