ß-Glactosidase from Bifidobacterium adolescentis DSM20083 prefers ß(1,4)-galactosides over lactose

S.W.A. Hinz; L.A.M. van den Broek; G. Beldman; J.P. Vincken

doi:10.1007/s00253-004-1745-9

ß-Glactosidase from Bifidobacterium adolescentis DSM20083 prefers ß(1,4)-galactosides over lactose

S.W.A. Hinz, L.A.M. van den Broek, G. Beldman, J.P. Vincken

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

65 Citations (Scopus)

Abstract

A -galactosidase gene (-Gal II) from Bifidobacterium adolescentis DSM 20083 was cloned into a pbluescript SK (–) vector and expressed in Escherichia coli. The recombinant enzyme was purified from the cell extract by anion-exchange and size-exclusion chromatography. -Gal II had a native molecular mass of 235 kDa and the subunits had a molecular mass of 81 kDa, indicating that -Gal II occurs as a trimer. The enzyme was classified as belonging to glycosyl hydrolase family 42. The optimal pH was 6.0 and the optimal temperature was 50°C, using p-nitrophenyl--d-galactopyranoside as a substrate. The Km and Vmax for Gal(1–4)Gal were 60 mM and 1,129 U/mg, respectively. The recombinant -Gal II was highly active towards Gal(1–4)Gal and Gal(1–4)Gal-containing oligosaccharides; only low activity was observed towards Gal(1–3)Gal, lactose, and Gal(1–3)GalOMe. No activity was found towards Gal(1–6)Gal, Gal(1–4)Man, Gal(1–4)Gal, Gal(1–3)Gal(1–4)Gal, cellobiose, maltose and sucrose. -Gal II was inhibited at high substrate concentrations (100 mg/ml) and no transglycosylation activity was found. At lower substrate concentrations (10 mg/ml) only low transglycosylation activity was found; the Gal/[Gal(1–4)]2Gal peak area ratio was 9:1.

Original language	English
Pages (from-to)	276-284
Journal	Applied Microbiology and Biotechnology
Volume	66
Issue number	3
DOIs	https://doi.org/10.1007/s00253-004-1745-9
Publication status	Published - 2004

Keywords

fermentation properties
oligosaccharides
purification
sequence
infantis
bifidum
system
classification
prediction
expression

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10.1007/s00253-004-1745-9

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@article{ccdc4ffec9704434ac6553623fcaebb6,

title = "{\ss}-Glactosidase from Bifidobacterium adolescentis DSM20083 prefers {\ss}(1,4)-galactosides over lactose",

abstract = "A -galactosidase gene (-Gal II) from Bifidobacterium adolescentis DSM 20083 was cloned into a pbluescript SK (–) vector and expressed in Escherichia coli. The recombinant enzyme was purified from the cell extract by anion-exchange and size-exclusion chromatography. -Gal II had a native molecular mass of 235 kDa and the subunits had a molecular mass of 81 kDa, indicating that -Gal II occurs as a trimer. The enzyme was classified as belonging to glycosyl hydrolase family 42. The optimal pH was 6.0 and the optimal temperature was 50°C, using p-nitrophenyl--d-galactopyranoside as a substrate. The Km and Vmax for Gal(1–4)Gal were 60 mM and 1,129 U/mg, respectively. The recombinant -Gal II was highly active towards Gal(1–4)Gal and Gal(1–4)Gal-containing oligosaccharides; only low activity was observed towards Gal(1–3)Gal, lactose, and Gal(1–3)GalOMe. No activity was found towards Gal(1–6)Gal, Gal(1–4)Man, Gal(1–4)Gal, Gal(1–3)Gal(1–4)Gal, cellobiose, maltose and sucrose. -Gal II was inhibited at high substrate concentrations (100 mg/ml) and no transglycosylation activity was found. At lower substrate concentrations (10 mg/ml) only low transglycosylation activity was found; the Gal/[Gal(1–4)]2Gal peak area ratio was 9:1.",

keywords = "fermentation properties, oligosaccharides, purification, sequence, infantis, bifidum, system, classification, prediction, expression",

author = "S.W.A. Hinz and {van den Broek}, L.A.M. and G. Beldman and J.P. Vincken",

year = "2004",

doi = "10.1007/s00253-004-1745-9",

language = "English",

volume = "66",

pages = "276--284",

journal = "Applied Microbiology and Biotechnology",

issn = "0175-7598",

publisher = "Springer",

number = "3",

}

TY - JOUR

T1 - ß-Glactosidase from Bifidobacterium adolescentis DSM20083 prefers ß(1,4)-galactosides over lactose

AU - Hinz, S.W.A.

AU - van den Broek, L.A.M.

AU - Beldman, G.

AU - Vincken, J.P.

PY - 2004

Y1 - 2004

N2 - A -galactosidase gene (-Gal II) from Bifidobacterium adolescentis DSM 20083 was cloned into a pbluescript SK (–) vector and expressed in Escherichia coli. The recombinant enzyme was purified from the cell extract by anion-exchange and size-exclusion chromatography. -Gal II had a native molecular mass of 235 kDa and the subunits had a molecular mass of 81 kDa, indicating that -Gal II occurs as a trimer. The enzyme was classified as belonging to glycosyl hydrolase family 42. The optimal pH was 6.0 and the optimal temperature was 50°C, using p-nitrophenyl--d-galactopyranoside as a substrate. The Km and Vmax for Gal(1–4)Gal were 60 mM and 1,129 U/mg, respectively. The recombinant -Gal II was highly active towards Gal(1–4)Gal and Gal(1–4)Gal-containing oligosaccharides; only low activity was observed towards Gal(1–3)Gal, lactose, and Gal(1–3)GalOMe. No activity was found towards Gal(1–6)Gal, Gal(1–4)Man, Gal(1–4)Gal, Gal(1–3)Gal(1–4)Gal, cellobiose, maltose and sucrose. -Gal II was inhibited at high substrate concentrations (100 mg/ml) and no transglycosylation activity was found. At lower substrate concentrations (10 mg/ml) only low transglycosylation activity was found; the Gal/[Gal(1–4)]2Gal peak area ratio was 9:1.

AB - A -galactosidase gene (-Gal II) from Bifidobacterium adolescentis DSM 20083 was cloned into a pbluescript SK (–) vector and expressed in Escherichia coli. The recombinant enzyme was purified from the cell extract by anion-exchange and size-exclusion chromatography. -Gal II had a native molecular mass of 235 kDa and the subunits had a molecular mass of 81 kDa, indicating that -Gal II occurs as a trimer. The enzyme was classified as belonging to glycosyl hydrolase family 42. The optimal pH was 6.0 and the optimal temperature was 50°C, using p-nitrophenyl--d-galactopyranoside as a substrate. The Km and Vmax for Gal(1–4)Gal were 60 mM and 1,129 U/mg, respectively. The recombinant -Gal II was highly active towards Gal(1–4)Gal and Gal(1–4)Gal-containing oligosaccharides; only low activity was observed towards Gal(1–3)Gal, lactose, and Gal(1–3)GalOMe. No activity was found towards Gal(1–6)Gal, Gal(1–4)Man, Gal(1–4)Gal, Gal(1–3)Gal(1–4)Gal, cellobiose, maltose and sucrose. -Gal II was inhibited at high substrate concentrations (100 mg/ml) and no transglycosylation activity was found. At lower substrate concentrations (10 mg/ml) only low transglycosylation activity was found; the Gal/[Gal(1–4)]2Gal peak area ratio was 9:1.

KW - fermentation properties

KW - oligosaccharides

KW - purification

KW - sequence

KW - infantis

KW - bifidum

KW - system

KW - classification

KW - prediction

KW - expression

U2 - 10.1007/s00253-004-1745-9

DO - 10.1007/s00253-004-1745-9

M3 - Article

SN - 0175-7598

VL - 66

SP - 276

EP - 284

JO - Applied Microbiology and Biotechnology

JF - Applied Microbiology and Biotechnology

IS - 3

ER -

ß-Glactosidase from Bifidobacterium adolescentis DSM20083 prefers ß(1,4)-galactosides over lactose

Abstract

Keywords

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