Purification and characterization of a protein from chicken gizzard, which inhibits actin polymerization.
An actin-polymerization-inhibiting protein, that occurs in crude preparations of vinculin from chicken gizzard, has been purified by DEAE-cellulose and carboxymethyl ion-exchange chromatography. According to sodium dodecyl sulfate (SDS)/polyacrylamide gel electrophoresis and to gel filtration the polymerization-inhibiting protein is heterogeneous and the molecular mass ranges from 20 kDa to 80 kDa. After treatment with acid the polymerization-inhibiting activity was found to migrate on a SDS/polyacrylamide gel as a single band of molecular mass about 32 kDa. The mechanism of the action of the polymerization-inhibiting protein on actin assembly was investigated by the effect on the kinetics of actin polymerization. The polymerization-inhibiting protein blocks elongation of actin filaments at substoichiometric ratios but does not nucleate actin filaments. The equilibrium constant for binding of the polymerization-inhibiting protein to the barbed end of an actin filament was estimated to be 2 X 10(6) M-1 in 100 mM KCl and 2 mM MgCl2, and 35 X 10(6) M-1 in 2 mM MgCl2.
D051339:Actin Depolymerizing Factors; D000199:Actins / Q000378:metabolism*; D000818:Animals; D001704:Biopolymers; D002645:Chickens; D002848:Chromatography, DEAE-Cellulose; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D051345:Destrin; D004591:Electrophoresis, Polyacrylamide Gel; D005895:Gizzard, Avian / Q000032:analysis*; D008840:Microfilament Proteins; D011485:Protein Binding; D011487:Protein Conformation; D011506:Proteins / Q000302:isolation & purification* / Q000502:physiology; D013050:Spectrometry, Fluorescence
D051339:Actin Depolymerizing Factors; D000199:Actins; D001704:Biopolymers; D051345:Destrin; D008840:Microfilament Proteins; D011506:Proteins