Kinetics of the interaction of a 41-kilodalton macrophage capping protein with actin: promotion of nucleation during prolongation of the lag period.
A 41-kilodalton macrophage capping protein (MCP) has been isolated which is capable of forming complexes with actin monomers in addition to capping the barbed ends of actin filaments (Southwick & DiNubile, 1986). The protein is calcium activated in a fully reversible manner. Using kinetic assays, we determined a capping constant, defined here as a modified Kd, of 1 nM and a Kd of 3-4 microM for MCP-actin monomer complex formation. MCP weakly nucleates actin polymerization: more than 0.5 microM MCP is necessary to shorten the lag period, and 1 microM MCP at an actin/MCP ratio of 10 reduces the average length of actin filaments to about 200 molecules per filament. We determined that the actin nucleus that survives MCP inactivation contains a minimum number of five actin molecules. These experiments also make a point with respect to the interpretation of the prolongation of the lag period. We directly demonstrate that in the presence of an actin binding protein a prolongation of the lag period can be associated with increased nucleation, contrary to the usual interpretation in the literature that it indicates no or decreased nucleation by the actin binding protein.
Young
C L
CL
Department of Medicine, University of Pennsylvania, Philadelphia 19104.
Southwick
F S
FS
Weber
A
A
D000177:Acrosome / Q000378:metabolism; D008841:Actin Cytoskeleton / Q000378:metabolism*; D051339:Actin Depolymerizing Factors; D000199:Actins / Q000378:metabolism*; D000818:Animals; D002118:Calcium / Q000494:pharmacology; D003599:Cytoskeleton / Q000378:metabolism*; D051345:Destrin; D005453:Fluorescence; D006737:Horseshoe Crabs; D007700:Kinetics; D008264:Macrophages / Q000378:metabolism*; D008297:Male; D008840:Microfilament Proteins / Q000378:metabolism*; D011108:Polymers
D051339:Actin Depolymerizing Factors; D000199:Actins; D051345:Destrin; D008840:Microfilament Proteins; D011108:Polymers; D002118:Calcium