Fungal Alpha-Galactosidase from Pseudobalsamia microspora Capable of Degrading Raffinose Family Oligosaccharides

An alpha-galactosidase was purified from Pseudobalsamia microspora (PMG) to 1224.1-fold with a specific activity of 11,274.5 units/mg by ion-exchange chromatography and gel filtration. PMG is a monomeric protein with a molecular mass of 62 kDa as determined by SDS-PAGE and by gel filtration. Chemica...

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Bibliographic Details
Published in:Applied biochemistry and biotechnology Vol. 176; no. 8; pp. 2157 - 2169
Main Authors: Yang, Dongxue, Tian, Guoting, Du, Fang, Zhao, Yongchang, Zhao, Liyan, Wang, Hexiang, Ng, Tzi Bun
Format: Journal Article
Language:English
Published: New York Springer US 01.08.2015
Springer Nature B.V
Subjects:
alpha-galactosidase
alpha-Galactosidase - chemistry
alpha-Galactosidase - isolation & purification
alpha-Galactosidase - metabolism
Amino Acid Sequence
Amino acids
Bacteria
Biochemistry
Biotechnology
cadmium
Chemistry
Chemistry and Materials Science
chromatography
Chromatography, Ion Exchange
Chromatography, Thin Layer
copper
Electrophoresis, Polyacrylamide Gel
essential amino acids
Feed industry
Filtration
Fungi
gel chromatography
Hydrogen-Ion Concentration
Hydrolysis
Indicators and Reagents
ion exchange chromatography
Ions
iron
Kinetics
Liquid chromatography
Mass spectrometry
mercury
Metals - pharmacology
Microspora
Microsporidia - enzymology
Microsporidia - isolation & purification
Molecular Sequence Data
Molecular Weight
Mushrooms
polyacrylamide gel electrophoresis
raffinose
Raffinose - metabolism
stachyose
Substrate Specificity - drug effects
tandem mass spectrometry
Temperature
Thin layer chromatography
Degradation
Purification and characterization
Chemical modification
Alpha-galactosidase
ISSN:0273-2289, 1559-0291, 1559-0291
Online Access:Get full text
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Summary:An alpha-galactosidase was purified from Pseudobalsamia microspora (PMG) to 1224.1-fold with a specific activity of 11,274.5 units/mg by ion-exchange chromatography and gel filtration. PMG is a monomeric protein with a molecular mass of 62 kDa as determined by SDS-PAGE and by gel filtration. Chemical modification using N-bromosuccinimide (NBS) resulted in a complete abrogation of the activity of PMG, suggesting that Trp is an amino acid essential to its activity. The activity was strongly inhibited by Hg²⁺, Cd²⁺, Cu²⁺, and Fe³⁺ ions. Three inner peptide sequences for PMG were obtained by liquid chromatography–tandem mass spectrometry (LC–MS–MS) analysis. When 4-nitrophenyl α-D-glucopyranoside (pNPGal) was used as substrate, the optimum pH and temperature of PMG were 5.0 and 55 °C, respectively. The Michaelis constant (K ₘ) value of the alpha-galactosidase on pNPGal was 0.29 mM, and the maximal velocity (V ₘₐₓ) was 0.97 μmol ml⁻¹ min⁻¹. Investigation by thin-layer chromatography (TLC) demonstrated its ability to hydrolyze raffinose and stachyose. Hence, it can be exploited in degradation of non-digestible oligosaccharides from food and feed industries.
Bibliography:http://dx.doi.org/10.1007/s12010-015-1705-0
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ISSN:0273-2289
1559-0291
1559-0291
DOI:10.1007/s12010-015-1705-0