Characterization and subunit structure of alpha-amylase inhibitors from seeds of two cultivars (Kintoki and Fukushirokintoki) of Phaseolus vulgaris L
2001
Sawada, S. (Mukogawa Women's Univ., Nishinomiya, Hyogo (Japan)) | Takeda, Y. | Kanamori, M. | Tashiro, M.
Two alpha-amylase inhibitors (KAI and FAI) were purified to apparent homogeneity from the respective seeds of Phaseolus vulgaris L.cv Kintoki and P.vulgaris L.cv Fukushirokintoki, and their properties and subunit structure were examined. Isoelectric points of KAI and FAI were 4.6 and 4.5, respectively, and each inhibitor was found to be a glycoprotein with a molecular weight of about 45000. Amino acid compositions of the inhibitors were similar to each other and characterized by no cysteine content. SDS-PAGE of each inhibitor showed at least four kinds of polypeptide bands corresponding to molecular weights between 14000 and 20000. On the other hand, size-exclusion HPLC indicated that each inhibitor was separated into two protein peaks, 1 and 2 under a denaturing condition. Amino acid compositions of 1 and 2 of KAI were very similar to those of 1 and 2 of FAI, respectively. N-terminal 20 amino acid sequences of 1 and 2 of KAI were the same as those of the correspondings of FAI. Comparison of 1 and 2 of each inhibitor exhibited that they were different from each other in their amino acid compositions and sequences. These findings suggest that each of KAI and FAI is composed of two protein subunits, which showed heterogeneity in molecular size due to different degrees of glycosylation.
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