Improving properties of a novel β-galactosidase from Lactobacillus plantarum by covalent immobilization

Rocio Benavente; Benevides C. Pessela; Jose Antonio Curiel; Blanca de las Rivas; Rosario Muñoz; Jose Manuel Guisán; Jose M. Mancheño; Alejandra Cardelle-Cobas; Ana I. Ruiz-Matute; Nieves Corzo

doi:10.3390/molecules20057874

Improving properties of a novel β-galactosidase from Lactobacillus plantarum by covalent immobilization

Rocio Benavente, Benevides C. Pessela^*, Jose Antonio Curiel, Blanca de las Rivas, Rosario Muñoz, Jose Manuel Guisán, Jose M. Mancheño, Alejandra Cardelle-Cobas, Ana I. Ruiz-Matute, Nieves Corzo

^*Corresponding author for this work

Research output: Contribution to journal › Article › peer-review

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Abstract

A novel β-galactosidase from Lactobacillus plantarum (LPG) was over-expressed in E. coli and purified via a single chromatographic step by using lowly activated IMAC (immobilized metal for affinity chromatography) supports. The pure enzyme exhibited a high hydrolytic activity of 491 IU/mL towards o-nitrophenyl β-D-galactopyranoside. This value was conserved in the presence of different divalent cations and was quite resistant to the inhibition effects of different carbohydrates. The pure multimeric enzyme was stabilized by multipoint and multisubunit covalent attachment on glyoxyl-agarose. The glyoxyl-LPG immobilized preparation was over 20-fold more stable than the soluble enzyme or the one-point CNBr-LPG immobilized preparation at 50°C. This β-galactosidase was successfully used in the hydrolysis of lactose and lactulose and formation of different oligosaccharides was detected. High production of galacto-oligosaccharides (35%) and oligosaccharides derived from lactulose (30%) was found and, for the first time, a new oligosaccharide derived from lactulose, tentatively identified as 3′-galactosyl lactulose, has been described.

Original language	English
Pages (from-to)	7874-7889
Number of pages	16
Journal	Molecules
Volume	20
Issue number	5
DOIs	https://doi.org/10.3390/molecules20057874
Publication status	Published - 1 May 2015

Keywords

Glyoxyl-agarose
Immobilization
Lactobacillus plantarum
Lactose
Lactulose
Oligosaccharides synthesis
β-galactosidase

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10.3390/molecules20057874Licence: CC BY

26871289Final published version, 1.36 MBLicence: CC BY

http://hdl.handle.net/10400.14/19934Licence: CC BY

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title = "Improving properties of a novel β-galactosidase from Lactobacillus plantarum by covalent immobilization",

abstract = "A novel β-galactosidase from Lactobacillus plantarum (LPG) was over-expressed in E. coli and purified via a single chromatographic step by using lowly activated IMAC (immobilized metal for affinity chromatography) supports. The pure enzyme exhibited a high hydrolytic activity of 491 IU/mL towards o-nitrophenyl β-D-galactopyranoside. This value was conserved in the presence of different divalent cations and was quite resistant to the inhibition effects of different carbohydrates. The pure multimeric enzyme was stabilized by multipoint and multisubunit covalent attachment on glyoxyl-agarose. The glyoxyl-LPG immobilized preparation was over 20-fold more stable than the soluble enzyme or the one-point CNBr-LPG immobilized preparation at 50°C. This β-galactosidase was successfully used in the hydrolysis of lactose and lactulose and formation of different oligosaccharides was detected. High production of galacto-oligosaccharides (35%) and oligosaccharides derived from lactulose (30%) was found and, for the first time, a new oligosaccharide derived from lactulose, tentatively identified as 3′-galactosyl lactulose, has been described.",

keywords = "Glyoxyl-agarose, Immobilization, Lactobacillus plantarum, Lactose, Lactulose, Oligosaccharides synthesis, β-galactosidase",

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T1 - Improving properties of a novel β-galactosidase from Lactobacillus plantarum by covalent immobilization

AU - Benavente, Rocio

AU - Pessela, Benevides C.

AU - Curiel, Jose Antonio

AU - Rivas, Blanca de las

AU - Muñoz, Rosario

AU - Guisán, Jose Manuel

AU - Mancheño, Jose M.

AU - Cardelle-Cobas, Alejandra

AU - Ruiz-Matute, Ana I.

AU - Corzo, Nieves

PY - 2015/5/1

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N2 - A novel β-galactosidase from Lactobacillus plantarum (LPG) was over-expressed in E. coli and purified via a single chromatographic step by using lowly activated IMAC (immobilized metal for affinity chromatography) supports. The pure enzyme exhibited a high hydrolytic activity of 491 IU/mL towards o-nitrophenyl β-D-galactopyranoside. This value was conserved in the presence of different divalent cations and was quite resistant to the inhibition effects of different carbohydrates. The pure multimeric enzyme was stabilized by multipoint and multisubunit covalent attachment on glyoxyl-agarose. The glyoxyl-LPG immobilized preparation was over 20-fold more stable than the soluble enzyme or the one-point CNBr-LPG immobilized preparation at 50°C. This β-galactosidase was successfully used in the hydrolysis of lactose and lactulose and formation of different oligosaccharides was detected. High production of galacto-oligosaccharides (35%) and oligosaccharides derived from lactulose (30%) was found and, for the first time, a new oligosaccharide derived from lactulose, tentatively identified as 3′-galactosyl lactulose, has been described.

AB - A novel β-galactosidase from Lactobacillus plantarum (LPG) was over-expressed in E. coli and purified via a single chromatographic step by using lowly activated IMAC (immobilized metal for affinity chromatography) supports. The pure enzyme exhibited a high hydrolytic activity of 491 IU/mL towards o-nitrophenyl β-D-galactopyranoside. This value was conserved in the presence of different divalent cations and was quite resistant to the inhibition effects of different carbohydrates. The pure multimeric enzyme was stabilized by multipoint and multisubunit covalent attachment on glyoxyl-agarose. The glyoxyl-LPG immobilized preparation was over 20-fold more stable than the soluble enzyme or the one-point CNBr-LPG immobilized preparation at 50°C. This β-galactosidase was successfully used in the hydrolysis of lactose and lactulose and formation of different oligosaccharides was detected. High production of galacto-oligosaccharides (35%) and oligosaccharides derived from lactulose (30%) was found and, for the first time, a new oligosaccharide derived from lactulose, tentatively identified as 3′-galactosyl lactulose, has been described.

KW - Glyoxyl-agarose

KW - Immobilization

KW - Lactobacillus plantarum

KW - Lactose

KW - Lactulose

KW - Oligosaccharides synthesis

KW - β-galactosidase

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Improving properties of a novel β-galactosidase from Lactobacillus plantarum by covalent immobilization

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