Nucleation of Actin Polymerization by Gelsolin

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Abstract
The time‐course of assembly of actin with gelsolin was measured by the fluorescence increase of a fluorescent label covalently linked to actin. The actin concentrations ranged from values far below the critical concentration to values above the critical concentration of the pointed ends of actin filaments. If the concentration of actin was in the range of the critical monomer concentration (0.64 μM), the time‐course of the concentration of actin assembled with gelsolin revealed a sigmoidal shape. At higher actin concentrations the time‐course of association of actin with gelsolin approximated an exponential curve. The measured time‐courses of assembly were quantitatively interpreted by kinetic rate equations. A poor fit was obtained if two actin molecules were assumed to bind to gelsolin to form a 1:2 gelsolin‐actin complex and subsequently further actin molecules were assumed to polymerize onto the 1:2 gelsolin‐actin complex toward the pointed end. A considerably better agreement between calculated and measured time‐courses was achieved if additional creation of actin filaments by fast fragmentation of newly formed actin filaments by not yet consumed gelsolin was assumed to occur. This suggests that both polymerization of actin onto gelsolin and fragmentation of actin filaments contribute to formation of new actin filaments by gelsolin. Furthermore it could be demonstrated that below the critical monomer concentration appreciable amounts of actin are incorporated into gelsolin‐actin oligomers.