Purification and characterization of isoforms of β-galactosidases in mung bean seedlings

Sing Chung Li, Jiahn Wern Han, Kuan Chung Chen, Ching San Chen

Research output: Contribution to journalArticle

52 Citations (Scopus)

Abstract

Five isoforms of β-galactosidase (EC 3.2.1.23), designated as β-galactosidases I-V, were isolated from five-day-old mung bean (Vigna radiata) seedlings. β-Galactosidases II and III were purified to electrophoretic homogeneity by a procedure involving acid precipitation, ammonium sulfate fractionation, chromatography on diethylaminoethyl-cellulose (DEAE-Cellulose) and con A-Sepharose, and chromatofocusing. β-Galactosidases I, II and III have the same molecular mass of 87 kDa, comprising two non-identical subunits with molecular masses of 38 and 48 kDa, while β-galactosidases IV and V have molecular masses of 45 and 73 kDa, respectively. All the enzymes were active against p-nitrophenyl-β-D-galactoside, and to a lesser extent, p-nitrophenyl-α-L-arabinoside and p-nitrophenyl-β-D-fucoside. The enzymes were inhibited by D-galactono-1,4-lactone, D-galactose, Hg2+, Ag+ and sodium dodecyl sulfate (SDS). β-Galactosidases I, II and III were shown to be competitively inhibited by either D-galactono-1, 4-lactone or D-galactose. Isoforms I, II and III have a common optimal pH of 3.6, while isoforms IV and V have pH optima at 3.8 and 4.0, respectively. Isoelectric points of isoforms I, II and III were 7.7, 7.5 and 7.3, respectively. Double immunodiffusion analysis indicated that β-galactosidases I, II, III and V are immunologically similar to each other, while β-galactosidase IV shares partially identical antigenic determinants with the other four isoforms. The purified β-galactosidases II and III were capable of releasing D-galactose residue from the hemicellulose fraction isolated from mung bean seeds.

Original languageEnglish
Pages (from-to)349-359
Number of pages11
JournalPhytochemistry
Volume57
Issue number3
DOIs
Publication statusPublished - Jun 2001
Externally publishedYes

Keywords

  • β-Galactosidase
  • Enzyme purification
  • Leguminosae
  • Natural substrates
  • Vigna radiata

ASJC Scopus subject areas

  • Plant Science
  • Biochemistry
  • Molecular Biology
  • Organic Chemistry
  • Drug Discovery

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