Purification and characterization of isoforms of β-galactosidases in mung bean seedlings
| Autor: | Sing Chung Li, Ching San Chen, Jiahn Wern Han, Kuan Chung Chen |
|---|---|
| Rok vydání: | 2001 |
| Předmět: |
Lactose
Plant Science Horticulture Biochemistry Isozyme Chromatography Affinity Chromatography DEAE-Cellulose Substrate Specificity Vigna chemistry.chemical_compound Enzyme Inhibitors Sodium dodecyl sulfate Molecular Biology chemistry.chemical_classification Plants Medicinal Chromatography Galactosidases biology Molecular mass Chromatofocusing Monosaccharides Fabaceae General Medicine beta-Galactosidase biology.organism_classification Isoenzymes Molecular Weight Kinetics Protein Subunits Isoelectric point Enzyme chemistry |
| Zdroj: | Phytochemistry. 57:349-359 |
| ISSN: | 0031-9422 |
| Popis: | Five isoforms of beta-galactosidase (EC 3.2.1.23), designated as beta-galactosidases I-V, were isolated from five-day-old mung bean (Vigna radiata) seedlings. Beta-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. Beta-galactosidases I, II and III have the same molecular mass of 87 kDa. comprising two nonidentical subunits with molecular masses of 38 and 48 kDa, while beta-galactosidases IV and V have molecular masses of 45 and 73 kDa, respectively. All the enzymes were active against p-nitrophenyl-beta-D-galactoside, and to a lesser extent, p-nitrophenyl-alpha-L-arabinoside and p-nitrophenyl-beta-D-fucoside. The enzymes were inhibited by D-galactono-1,4-lactone, D-galactose, Hg2+, Ag+ and sodium dodecyl sulfate (SDS). Beta-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 beta-galactosidases I, II, III and V are immunologically similar to each other, while beta-galactosidase IV shares partially identical antigenic determinants with the other four isoforms. The purified beta-galactosidases II and III were capable of releasing D-galactose residue from the hemicellulose fraction isolated from mung bean seeds. |
| Databáze: | OpenAIRE |
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