Purification of secreted α-amylases by immunoaffinity chromatography with cross-reactive antibody

Abstract

Two isozymes of rice α-amylases expressed and secreted by recombinant yeast were purified by immunoaffinity chromatography by using cross-reactive antibody. Antibodies raised against partially purified barley α-amylase adsorbed rice α-amylases in fermentation broth by a cross-reaction. By use of these antibodies as ligands, rice α-amylases were concentrated and purified to a high degree in one-step immunoaffinity chromatography. Because of the differences in the contaminating impurities between the barley α-amylase (antigen) from barley malt and rice α-amylases (target protein) secreted from yeast, the high purity of eluted α-amylases was attained without the use of highly purified antigen for immunization. Utilization of cross-reactive antibodies in immunoaffinity chromatography is useful for the purification of recombinant proteins in the absence of a sufficient amount and high enough purity of the target proteins to be purified.