Molecular characterization of DSR-E, an alpha-1,2 linkage-synthesizing dextransucrase with two catalytic domains.
A novel Leuconostoc mesenteroides NRRL B-1299 dextransucrase gene, dsrE, was isolated, sequenced, and cloned in Escherichia coli, and the recombinant enzyme was shown to be an original glucansucrase which catalyses the synthesis of alpha-1,6 and alpha-1,2 linkages. The nucleotide sequence of the dsrE gene consists of an open reading frame of 8,508 bp coding for a 2,835-amino-acid protein with a molecular mass of 313,267 Da. This is twice the average mass of the glucosyltransferases (GTFs) known so far, which is consistent with the presence of an additional catalytic domain located at the carboxy terminus of the protein and of a central glucan-binding domain, which is also significantly longer than in other glucansucrases. From sequence comparison with family 70 and alpha-amylase enzymes, crucial amino acids involved in the catalytic mechanism were identified, and several original sequences located at some highly conserved regions in GTFs were observed in the second catalytic domain.
Bozonnet
Sophie
S
Centre de Bioingéniérie Gilbert Durand, UMR CNRS 5504, UMR INRA 792, DGBA INSA, 31077 Toulouse Cedex 04, France.
Dols-Laffargue
Marguerite
M
Fabre
Emeline
E
Pizzut
Sandra
S
Remaud-Simeon
Magali
M
Monsan
Pierre
P
Willemot
René-Marc
RM
D000595:Amino Acid Sequence; D020134:Catalytic Domain / Q000235:genetics*; D003001:Cloning, Molecular; D003911:Dextrans / Q000737:chemistry* / Q000378:metabolism; D005964:Glucosyltransferases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D007934:Leuconostoc / Q000201:enzymology* / Q000235:genetics; D008969:Molecular Sequence Data; D009844:Oligosaccharides / Q000737:chemistry* / Q000378:metabolism; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA
D003911:Dextrans; D009844:Oligosaccharides; D005964:Glucosyltransferases; C020025:dextransucrase