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Isolation and characterization of alpha-amylases from endosperm of germinating maize

Author:
Warner, D.A., Grove, M.J., Knutson, C.A.
Source:
Cereal chemistry 1991 v.68 no.4 pp. 383
ISSN:
0009-0352
Subject:
Zea mays, alpha-amylase, purification, enzyme activity, endosperm, seed germination, starch, hydrolysis
Abstract:
Amylases from germinating maize (cv. B73) were fractionated by affinity chromatography, anion exchange chromatography, and chromatofocusing. Two groups of amylase enzymes were separated by affinity chromatography. About one half of the total amylase activity was bound on a cycloheptaamylose-Sepharose 6B column. Bound proteins were fractionated by anion exchange into four major .alpha.-amylase fractions, then further separated by chromatofocusing into eight fractions with apparent isoelectric point (pI) values ranging from 5.70 to 4.06. All affinity-bound fractions were confirmed as .alpha.-amylases by their action of .beta.-limit dextrin. The affinity-bound .alpha.-amylases with lowest and highest pI values produced reaction products from soluble starch containing large amounts of dextrins with degrees of polymerization (DP) 2 and 6, with lesser amounts of intermediate oligosaccharides. Intermediate pI fractions produced primarily DP2, large amounts of DP3-5, and little DP6. Enzymes not bound by cycloheptaamylose affinity chromatography were purified by hydroxylapatite chromatography, then resolved by chromatofocusing into four subgroups of .alpha.-amylase, plus .beta.-amylase. Among the affinity-unbound fractions, the lowest pI .alpha.-amylase had a unique action pattern, producing primarily DP7 and 8 oligosaccharides after exhaustive hydrolysis of soluble starch.
Agid:
24404
Handle:
10113/24404