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PURIFICATION AND CHARACTERIZATION OF SOY COTYLEDON β‐GLUCOSIDASE
Author(s) -
SANTOS R.F.,
OLIVEIRA C.F.,
VARÉA G.S.,
ORRADI DA SILVA M.L.C.,
IDA E.I.,
MANDARINO J.M.G.,
CARRÃOPANIZZI M.C.,
RIBEIRO M.L.L.
Publication year - 2013
Publication title -
journal of food biochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.507
H-Index - 47
eISSN - 1745-4514
pISSN - 0145-8884
DOI - 10.1111/j.1745-4514.2011.00632.x
Subject(s) - chemistry , chromatography , sephadex , daidzin , hydrolysis , aglycone , size exclusion chromatography , tricine , gel permeation chromatography , biochemistry , enzyme , glycoside , daidzein , organic chemistry , genistein , polymer , medicine
β‐Glucosidase F 42 of soy cotyledons was purified by ammonium sulfate fractionation, ion‐exchange chromatography (CM‐Sephadex‐C‐50, Sigma, St. Louis, MO) and gel filtration (Sephadex G‐100, Sigma). The enzyme was purified 111.8‐fold relative to its concentration in the crude extract. It had an apparent molecular mass of 53 kDa in gel filtration experiments and produced a 33‐kDa band in sodium dodecyl sulfate–polyacrylamide gel electrophoresis, suggesting that it is dimeric. The purified β‐glucosidase F 42 was characterized as a glycoprotein after the identification of fucose, galactosamine and glucosamine by high‐pressure anion‐exchange chromatography–pulsed amperometric detector. Its highest activity was observed at pH 5.0 and 45C, and it was stable for up to 4 days at 25C. The K m of the enzyme was 0.12 mM p ‐nitrophenyl‐β‐ d ‐glucopyranoside. β‐Glucosidase F 42 showed specificity for different substrates, and its activity was inhibited by 1 mM HgCl 2 , 10 mM glucono‐δ‐lactone or 150 mM glucose and increased by 10 mM MnCl 2 . PRACTICAL APPLICATIONS β‐Glucosidase is an enzyme that hydrolyzes β‐glucosidic bonds to liberate glucose and hydrolyzes isoflavones to release aglycones. Soy aglycones have been broadly investigated because of their biological activity in the prevention and treatment of some chronic diseases. Soy β‐glucosidase can be used in the food industry to alter soy isoflavones for the production of functional foods that are rich in aglycone isoflavones. Therefore, it was an established method of purification of the enzyme that has great biotechnological potential.