title: An Isoelectric Separation of Soybean β-Amylase Isoforms and Their Enzymic Characteristics.
abstract: Authentic soybean β-amylase preparation, purified to homogeneity as judged by SDS-PAGE by using an affinity purification step, was composed of four pI-differing isoforms. By chromatofocusing, these isoforms were separated into three fractions, designated as fractions 1-3 in the order of elution. Fraction 1 contained two isoforms having the same molecular mass (55,989 Da), as measured by mass spectrometric analysis, with different pIs, 5.32 (Isoform I) and 5.22 (Isoform II). Fraction 2 showed a single isoform having a molecular mass of 55,994 Da and having a pI of 5.09. This component, named Isoform III, existed rather in excess in a mixture of the authentic enzyme isoforms. The remainder (fraction 3) also contained a single component (Isoform IV) which has a molecular mass of 56,310 Da with a pI of 4.97. Chemical analyses indicated that the N-termini and the C-terminal tripeptides of four pI-separated isoforms mentioned are similar to one another, and are blocked and are NH2-Val-Asp-Gly-COOH, respectively. Moreover, enzymic properties involving specific activity and the value of kcat/Km for the above three fractions are almost the same, and also agreed completely with those of an unfractionated authentic β-amylase preparation.
authors.affiliation: a Genetic Engineering Laboratory, National Food Research Institute, Ministry of Agriculture, Forestry and Fisheries.
authors.forename: Y
authors.initials: Y
authors.lastname: Wang
authors.affiliation: a Genetic Engineering Laboratory, National Food Research Institute, Ministry of Agriculture, Forestry and Fisheries.
authors.forename: M
authors.initials: M
authors.lastname: Arahira
authors.affiliation: a Genetic Engineering Laboratory, National Food Research Institute, Ministry of Agriculture, Forestry and Fisheries.
authors.forename: C
authors.initials: C
authors.lastname: Fukazawa
keywords: chromatofocusing; kinetic analyses; mass spectrometric analyses; pI-heterogeneity; soybean β-amylase