The industrially important polysaccharide alginate is composed of
the two sugar monomers beta-D-mannuronic acid (M) and its epimer
alpha-L-guluronic acid (G). In the bacterium Azotobacter vinelandii,
the G residues originate from a polymer-level reaction catalyzed
by one periplasmic and at least five secreted mannuronan C-5-epimerases.
The secreted enzymes are composed of repeats of two protein modules
designated A (385 amino acids) and R (153 amino acids). The modular
structure of one of the epimerases, AlgE1, is A1R1R2R3A2R4. This
enzyme has two catalytic sites for epimerization, each site introducing
a different G distribution pattern, and in this article we report
the DNA-level construction of a variety of truncated forms of the
enzyme. Analyses of the properties of the corresponding proteins
showed that an A module alone is sufficient for epimerization and
that A1 catalyzed the formation of contiguous stretches of G residues
in the polymer, while A2 introduces single G residues. These differences
are predicted to strongly affect the physical and immunological properties
of the reaction product. The epimerization reaction is Ca2+ dependent,
and direct binding studies showed that both the A and R modules bind
this cation. The R modules appeared to reduce the Ca2+ concentration
needed for full activity and also stimulated the reaction rate when
positioned both N and C terminally.
UNIGEN Center for Molecular Biology and Department of Biotechnology,
Norwegian University of Technology and Science, N-7489 Trondheim,
Norway. Helga.Ertesvag@unigen.ntnu.no
%0 Journal Article
%1 Ertesvag1999
%A Ertesvag, H.
%A Valla, S.
%D 1999
%J J. Bacteriol.
%K & ; Acid Acids/analysis/metabolism Alginates/chemistry/metabolism Amino Azotobacter Binding Calcium/*metabolism/pharmacology Carbohydrate Catalysis/drug Catalytic Cations/metabolism/pharmacology Data Deletion Domain Epimerases/chemistry/genetics/isolation Escherichia Gov't Hexuronic Kinetics Magnetic Molecular Non-U.S. Plasmids/genetics Proteins/biosynthesis/chemistry/isolation Recombinant Relationship Research Resonance Sequence Sites Spectroscopy Structure-Activity Support, coli/genetics effects purification/*metabolism purification/metabolism vinelandii/*enzymology/genetics/metabolism
%N 10
%P 3033-8
%T The A modules of the Azotobacter vinelandii mannuronan-C-5-epimerase
AlgE1 are sufficient for both epimerization and binding of Ca2+.
%U http://eutils.ncbi.nlm.nih.gov/entrez/eutils/elink.fcgi?cmd=prlinks&dbfrom=pubmed&retmode=ref&id=10322003
%V 181
%X The industrially important polysaccharide alginate is composed of
the two sugar monomers beta-D-mannuronic acid (M) and its epimer
alpha-L-guluronic acid (G). In the bacterium Azotobacter vinelandii,
the G residues originate from a polymer-level reaction catalyzed
by one periplasmic and at least five secreted mannuronan C-5-epimerases.
The secreted enzymes are composed of repeats of two protein modules
designated A (385 amino acids) and R (153 amino acids). The modular
structure of one of the epimerases, AlgE1, is A1R1R2R3A2R4. This
enzyme has two catalytic sites for epimerization, each site introducing
a different G distribution pattern, and in this article we report
the DNA-level construction of a variety of truncated forms of the
enzyme. Analyses of the properties of the corresponding proteins
showed that an A module alone is sufficient for epimerization and
that A1 catalyzed the formation of contiguous stretches of G residues
in the polymer, while A2 introduces single G residues. These differences
are predicted to strongly affect the physical and immunological properties
of the reaction product. The epimerization reaction is Ca2+ dependent,
and direct binding studies showed that both the A and R modules bind
this cation. The R modules appeared to reduce the Ca2+ concentration
needed for full activity and also stimulated the reaction rate when
positioned both N and C terminally.
@article{Ertesvag1999,
__markedentry = {[phpts:6]},
abstract = {The industrially important polysaccharide alginate is composed of
the two sugar monomers beta-D-mannuronic acid (M) and its epimer
alpha-L-guluronic acid (G). In the bacterium Azotobacter vinelandii,
the G residues originate from a polymer-level reaction catalyzed
by one periplasmic and at least five secreted mannuronan C-5-epimerases.
The secreted enzymes are composed of repeats of two protein modules
designated A (385 amino acids) and R (153 amino acids). The modular
structure of one of the epimerases, AlgE1, is A1R1R2R3A2R4. This
enzyme has two catalytic sites for epimerization, each site introducing
a different G distribution pattern, and in this article we report
the DNA-level construction of a variety of truncated forms of the
enzyme. Analyses of the properties of the corresponding proteins
showed that an A module alone is sufficient for epimerization and
that A1 catalyzed the formation of contiguous stretches of G residues
in the polymer, while A2 introduces single G residues. These differences
are predicted to strongly affect the physical and immunological properties
of the reaction product. The epimerization reaction is Ca2+ dependent,
and direct binding studies showed that both the A and R modules bind
this cation. The R modules appeared to reduce the Ca2+ concentration
needed for full activity and also stimulated the reaction rate when
positioned both N and C terminally.},
added-at = {2011-11-04T13:47:04.000+0100},
author = {Ertesvag, H. and Valla, S.},
authoraddress = {UNIGEN Center for Molecular Biology and Department of Biotechnology,
Norwegian University of Technology and Science, N-7489 Trondheim,
Norway. Helga.Ertesvag@unigen.ntnu.no},
biburl = {https://www.bibsonomy.org/bibtex/2b597d60db7ba0f29f3f8047430301a91/pawelsikorski},
interhash = {fb5ca394d5818515b9bc35596d703206},
intrahash = {b597d60db7ba0f29f3f8047430301a91},
journal = {J. Bacteriol.},
keywords = {& ; Acid Acids/analysis/metabolism Alginates/chemistry/metabolism Amino Azotobacter Binding Calcium/*metabolism/pharmacology Carbohydrate Catalysis/drug Catalytic Cations/metabolism/pharmacology Data Deletion Domain Epimerases/chemistry/genetics/isolation Escherichia Gov't Hexuronic Kinetics Magnetic Molecular Non-U.S. Plasmids/genetics Proteins/biosynthesis/chemistry/isolation Recombinant Relationship Research Resonance Sequence Sites Spectroscopy Structure-Activity Support, coli/genetics effects purification/*metabolism purification/metabolism vinelandii/*enzymology/genetics/metabolism},
language = {eng},
medline-da = {19990617},
medline-dcom = {19990617},
medline-edat = {1999/05/13},
medline-fau = {Ertesvag, H ; Valla, S},
medline-is = {0021-9193 (Print)},
medline-jid = {2985120R},
medline-jt = {Journal of bacteriology.},
medline-lr = {20060501},
medline-mhda = {1999/05/13 00:01},
medline-own = {NLM},
medline-pl = {UNITED STATES},
medline-pmid = {10322003},
medline-pst = {ppublish},
medline-pt = {Journal Article},
medline-pubm = {Print},
medline-rn = {0 (Alginates) ; 0 (Cations) ; 0 (Hexuronic Acids) ; 0 (Recombinant
Proteins) ; 15769-56-9 (guluronic acid) ; 6814-36-4 (mannuronic acid)
; 7440-70-2 (Calcium) ; EC 5.1.3 (Carbohydrate Epimerases)},
medline-sb = {IM},
medline-so = {J Bacteriol. 1999 May;181(10):3033-8.},
medline-stat = {MEDLINE},
number = 10,
owner = {phpts},
pages = {3033-8},
timestamp = {2011-11-04T13:47:10.000+0100},
title = {The A modules of the Azotobacter vinelandii mannuronan-C-5-epimerase
AlgE1 are sufficient for both epimerization and binding of Ca2+.},
url = {http://eutils.ncbi.nlm.nih.gov/entrez/eutils/elink.fcgi?cmd=prlinks\&dbfrom=pubmed\&retmode=ref\&id=10322003},
volume = 181,
year = 1999
}