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Basic characterization and partial purification of β ‐glucosidase from cell‐free extracts of Oenococcus oeni ST81
Author(s) -
Mesas J.M.,
Rodríguez M.C.,
Alegre M.T.
Publication year - 2012
Publication title -
letters in applied microbiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.698
H-Index - 110
eISSN - 1472-765X
pISSN - 0266-8254
DOI - 10.1111/j.1472-765x.2012.03285.x
Subject(s) - oenococcus oeni , malolactic fermentation , biochemistry , lactobacillus brevis , lactobacillus plantarum , fructose , wine , periplasmic space , enzyme , chemistry , enzyme assay , sorbitol , ammonium sulfate precipitation , biology , escherichia coli , chromatography , lactic acid , bacteria , food science , size exclusion chromatography , genetics , gene
Aims: This study was designed to characterize a β‐glucosidase of Oenococcus oeni ST81, a strain isolated from a Spanish wine of the origin appellation Ribeira Sacra. Methods and Results: The β‐glucosidase of O. oeni ST81 seems to have a periplasmic localization into the cells. This activity was strongly inhibited by gluconic acid, partially inhibited by glucose and not inhibited by fructose, lactate, malate, mannitol or sorbitol. Ethanol increased the activity of this enzyme up to 147%. Among the several metal ions assayed, only Fe 2+ (10 mmol l −1 ) and Cu 2+ (5 mmol l −1 ) exhibited a partial inhibitory effect (40%). This enzyme was partially purified using a combination of ammonium sulfate precipitation and chromatographic methods. The single peak because of β‐glucosidase in all chromatographic columns indicates the presence of a single enzyme with an estimated molecular mass of 140 kDa. The calculated K m and V max values for 4‐nitrophenyl‐β‐ d ‐glucopyranoside were 0·38 mmol l −1 and 5·21 nmol min −1 , respectively. The enzyme was stable at pH 5·0 with a value of t 1/2 = 50 days for the crude extract. Conclusions: The β‐glucosidase of O. oeni ST81 is substantially different from those characterized from other wine‐related lactic acid bacteria (LAB), such as Lactobacillus plantarum and Lactobacillus brevis ; however, it appears to be closely related to a β‐glucosidase from O. oeni ATCC BAA‐1163 cloned into Escherichia coli . The periplasmic localization of the enzyme together with its high tolerance to ethanol and fructose, the low inhibitory effect of some wine‐related compounds on the enzymatic activity and long‐term stability of the enzyme could be of interest for winemaking. Significance and Impact of the Study: Information regarding a β‐glucosidase from O. oeni ST81 is presented. Although the release of aroma compounds by LAB has been demonstrated, little information exists concerning the responsible enzymes. To our knowledge, this study contains the first characterization of a native β‐glucosidase purified from crude extracts of O. oeni ST81.