PURIFICATION AND CHARACTERIZATION OF THE ACID-SOLUBLE 26-KDA POLYPEPTIDE FROM SOYBEAN SEEDS

Whey proteins from soybean seeds of Japanese varieties were analyzed b y SDS-polyacrylamide gel electrophoresis (SDS-PAGE), Among 11 varietie s of soybean, three green and one black soybeans Lacked a 26-kDa band that was found in all yellow soybeans, In this paper, the 26-kDa prote in was named AS26k (acid soluble 26-kDa protein) temporarily, The AS26 k protein was purified from Glycine max cv, Nattosyoryu, which is yell ow soybean, through four purification steps: 30-35% saturated ammonium sulfate fractionation, ion exchange chromatography on S Sepharose Fas t Flow, gel filtration on Sephadex G-100, and hydrophobic chromatograp hy on phenyl Sepharose CL-4B, Purified AS26k was cleaved with V8 prote inase from Staphylococcus aureus or CNBr. The cleaved polypeptide cont ained two typical dehydrin motif sequences: DEYGNPV and (M)DKIKEKLPG, and a 19 amino acids sequence similar to a pea dehydrin, Native AS26k had a molecular mass of 32 kDa on gel filtration and a pi of 7.2 on tw o-dimensional PAGE, Similarly to other dehydrins and late embryogenesi s abundant (LEA) proteins, AS26k was rich in hydrophilic amino acids, and highly heat stable. These results showed that AS26k was a dehydrin , a group II LEA protein in soybean seeds.