RAPID AND EFFICIENT PURIFICATION OF ACTIN FROM NONMUSCLE SOURCES

The need for biochemical quantities of nonmuscle actin has been increa sed by observations that actin isoform composition of a cell influence s the cell's motile and structural properties. In addition, the number of actin binding proteins that exhibit different binding interactions with beta- and gamma-actin compared to oi-actin from skeletal muscle is growing. We report a procedure designed to purify actin from nonmus cle tissues employing extraction of monomeric actin from tissues with high concentrations of Tris, chromatography on DE-53 cellulose, and af finity chromatography of DNase I-agarose. The preparation is easy to p erform and yields quantities of nonmuscle actin sufficient for biochem ical and cell biological assays. Actin from bovine erythrocytes and fr om brains of adult and embryonic chickens was obtained using this meth od. which can be readily used with other sources of tissue. Coomassie- Blue-stained SDS gels of the purified actin show no contaminants; capp ing protein, a common contaminant of actin preparations, is absent by immunoblotting. This method for purifying nonmuscle actin will be usef ul to investigate functional differences in the biology of actin isofo rms or their regulating proteins. (C) 1998 Wiley-Liss, Inc.