Mutational analysis of the role of calcium ions in the Lactobacillus reuteri strain 121 fructosyltransferase (levansucrase and inulosucrase) enzymes

L.K. Ozimek; G.J.W. Euverink; M.J.E.C. van der Maarel; L. Dijkhuizen

doi:10.1016/j.febslet.2004.11.113

Mutational analysis of the role of calcium ions in the Lactobacillus reuteri strain 121 fructosyltransferase (levansucrase and inulosucrase) enzymes

L.K. Ozimek, G.J.W. Euverink, M.J.E.C. van der Maarel, L. Dijkhuizen

Groningen Biomolecular Sciences and Biotechnology Institute

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Abstract

Bacterial fructosyltransferase enzymes belonging to glycoside hydrolase family 68 (GH68) are not known to require a metal cofactor. Here, we show that Ca2+ ions play an important structural role in the Lactobacillus reuteri 121 levansucrase (Lev) and inulosucrase (Inu) enzymes. Analysis of the Bacillus subtilis Lev 3D structure has provided evidence for the presence of a bound metal ion, most likely Ca2+. Characterization of site-directed mutants in the putative Ca2+ ion-binding sites of Lb. reuteri Lev and Inu revealed that the Inu Asp520 and Lev Asp500 residues play an important role in Ca2+ binding. Sequence alignments of family GH68 proteins showed that this Ca2+ ion-binding site is (largely) present only in proteins of Gram-positive origin.

Original language	English
Pages (from-to)	1124-1128
Number of pages	5
Journal	FEBS Letters
Volume	579
Issue number	5
DOIs	https://doi.org/10.1016/j.febslet.2004.11.113
Publication status	Published - 14-Feb-2005

Keywords

Calcium ion
Lactobacillus reuteri
Mutagenesis
Inulosucrase
Fructosyltransferase
Levansucrase

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title = "Mutational analysis of the role of calcium ions in the Lactobacillus reuteri strain 121 fructosyltransferase (levansucrase and inulosucrase) enzymes",

abstract = "Bacterial fructosyltransferase enzymes belonging to glycoside hydrolase family 68 (GH68) are not known to require a metal cofactor. Here, we show that Ca2+ ions play an important structural role in the Lactobacillus reuteri 121 levansucrase (Lev) and inulosucrase (Inu) enzymes. Analysis of the Bacillus subtilis Lev 3D structure has provided evidence for the presence of a bound metal ion, most likely Ca2+. Characterization of site-directed mutants in the putative Ca2+ ion-binding sites of Lb. reuteri Lev and Inu revealed that the Inu Asp520 and Lev Asp500 residues play an important role in Ca2+ binding. Sequence alignments of family GH68 proteins showed that this Ca2+ ion-binding site is (largely) present only in proteins of Gram-positive origin.",

keywords = "Calcium ion, Lactobacillus reuteri, Mutagenesis, Inulosucrase, Fructosyltransferase, Levansucrase",

author = "L.K. Ozimek and G.J.W. Euverink and Maarel, {M.J.E.C. van der} and L. Dijkhuizen",

note = "Relation: http://www.rug.nl/research/gbb/ date_submitted:2007 Rights: University of Groningen, Groningen Biomolecular Sciences and Biotechnology Institute",

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AU - Ozimek, L.K.

AU - Euverink, G.J.W.

AU - Maarel, M.J.E.C. van der

AU - Dijkhuizen, L.

N1 - Relation: http://www.rug.nl/research/gbb/ date_submitted:2007 Rights: University of Groningen, Groningen Biomolecular Sciences and Biotechnology Institute

PY - 2005/2/14

Y1 - 2005/2/14

N2 - Bacterial fructosyltransferase enzymes belonging to glycoside hydrolase family 68 (GH68) are not known to require a metal cofactor. Here, we show that Ca2+ ions play an important structural role in the Lactobacillus reuteri 121 levansucrase (Lev) and inulosucrase (Inu) enzymes. Analysis of the Bacillus subtilis Lev 3D structure has provided evidence for the presence of a bound metal ion, most likely Ca2+. Characterization of site-directed mutants in the putative Ca2+ ion-binding sites of Lb. reuteri Lev and Inu revealed that the Inu Asp520 and Lev Asp500 residues play an important role in Ca2+ binding. Sequence alignments of family GH68 proteins showed that this Ca2+ ion-binding site is (largely) present only in proteins of Gram-positive origin.

AB - Bacterial fructosyltransferase enzymes belonging to glycoside hydrolase family 68 (GH68) are not known to require a metal cofactor. Here, we show that Ca2+ ions play an important structural role in the Lactobacillus reuteri 121 levansucrase (Lev) and inulosucrase (Inu) enzymes. Analysis of the Bacillus subtilis Lev 3D structure has provided evidence for the presence of a bound metal ion, most likely Ca2+. Characterization of site-directed mutants in the putative Ca2+ ion-binding sites of Lb. reuteri Lev and Inu revealed that the Inu Asp520 and Lev Asp500 residues play an important role in Ca2+ binding. Sequence alignments of family GH68 proteins showed that this Ca2+ ion-binding site is (largely) present only in proteins of Gram-positive origin.

KW - Calcium ion

KW - Lactobacillus reuteri

KW - Mutagenesis

KW - Inulosucrase

KW - Fructosyltransferase

KW - Levansucrase

U2 - 10.1016/j.febslet.2004.11.113

DO - 10.1016/j.febslet.2004.11.113

M3 - Article

SN - 0014-5793

VL - 579

SP - 1124

EP - 1128

JO - FEBS Letters

JF - FEBS Letters

IS - 5

ER -

Mutational analysis of the role of calcium ions in the Lactobacillus reuteri strain 121 fructosyltransferase (levansucrase and inulosucrase) enzymes

Abstract

Keywords

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