IDENTIFICATION OF AN ETHANOL-SOLUBLE PROTEIN AS BETA-AMYLASE AND ITS PURIFICATION FROM SOYBEAN SEEDS

In the 60% ethanol extract of soybean seeds, a prominent protein band was visible after polyacrylamide gel electrophoresis, which had a mole cular weight of about 55 x 10(3) M(r). This protein was purified to ho mogeneity by buffered ethanol extraction and preparatory gel electroph oresis. Since the N-terminus was apparently blocked, the protein was c leaved with cyanogen bromide and the largest fragment was isolated and a partial sequence determined. The sequence of the 27 N-terminal amin o acid residues matched a published soybean beta-amylase peptide seque nce. In addition, the purified protein had a high specific activity fo r beta-amylase and was not a glycoprotein. Furthermore, the partial se quence (106 nucleotides) of a cDNA clone, isolated from a soybean seed cDNA library by antibody screening, matched the cDNA sequence of soyb ean beta-amylase except for one base. Therefore, the ethanol-soluble p rotein was identified as beta-amylase. The enzyme was purified to homo geneity using a two-step purification procedure with a yield of over 5 0%.