Molecular characterization and expression analysis of the glucansucrase DSRWC from Weissella cibaria synthesizing a alpha(1-->6) glucan.
Weissella cibaria isolated from human saliva produces a soluble glucan that predominantly has alpha-1,6-glucosidic type linkages. Using degenerated primers that were selected based on the amino acid sequences of conserved regions from known glucansucrases, a single 2.7-kb fragment was isolated. In subsequent steps, a 4969-bp product was obtained using inverse PCR. The coding region for the glucansucrase gene (dsrWC) consisted of a 4419-bp ORF that encoded a 1472-amino acid protein with a calculated molecular mass of 161.998 Da. The produced DSRWC glucansucrases exhibited similarity with the enzymes of the glucosylhydrolase family 70, which includes the Lactobacillus fermentum glucansucrase. The expressed recombinant DSRWC (rDSRWC) synthesized oligosaccharides in the presence of maltose or isomaltose as an acceptor and the synthesized products included alpha-1,6-linked glucosyl residues in addition to the maltosyl or isomaltosyl residue. rDSRWC synthesized water-soluble polymers using sucrose as substrate. According to the (13)C-nuclear magnetic resonance analysis, the polymer that was synthesized by rDSRWC was a linear dextran, which formed predominately alpha-1,6-glucosidic linkages. This is the first report on the molecular characterization of glucansucrase from a W. cibaria strain.
Kang
Hee-Kyoung
HK
The Research Institute for Catalysis, Chonnam National University, Gwang-Ju, South Korea.
Oh
Jong-Suk
JS
Kim
Doman
D
D000595:Amino Acid Sequence; D004269:DNA, Bacterial / Q000737:chemistry / Q000235:genetics; D004795:Enzyme Stability; D005936:Glucans / Q000378:metabolism*; D016695:Glycosyltransferases / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics*; D006094:Gram-Positive Bacteria / Q000201:enzymology* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007534:Isomaltose / Q000378:metabolism; D009682:Magnetic Resonance Spectroscopy; D008320:Maltose / Q000378:metabolism; D008969:Molecular Sequence Data; D008970:Molecular Weight; D016366:Open Reading Frames; D016133:Polymerase Chain Reaction / Q000379:methods; D012463:Saliva / Q000382:microbiology; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D013696:Temperature
D004269:DNA, Bacterial; D005936:Glucans; C504992:alpha-1-6-glucan; D007534:Isomaltose; D008320:Maltose; D016695:Glycosyltransferases; C033837:alternansucrase