Journal of Microbiology and Biotechnology

The Korean Society for Microbiology and Biotechnology (한국미생물·생명공학회)
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Cloning and Sequencing of the ${\alpha}-1{\rightarrow}6$ Dextransurcrase Gene from Leuconostoc mensenteroides B-742CB

  • Kim, Ho-Sang (The Research Center for New Bio-Materials in Agriculture, Seoul National University) ;
  • Kim, Do-Man (Department of Biochemical Engineering and Research Institute for Catalysis, Chonnam National University) ;
  • Ryu, Hwa-Ja (Department of Fine Chemical Engineering, Chonnam National University) ;
  • Robyt, John-F. (Lagoratory of Carbohydrate Chemistry and Enzymology, Iowa State University)
  • Published : 2000.08.01
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Abstract

A dextransucrase gene (dsrB742) that expresses a dextransucrase to synthesize mostly ${\alpha}-1{\rightarrow}6$ linked dextran with a low amount (3-5%) of ${\alpha}-1{\rightarrow}3$ branching was cloned and sequenced from Leuconostoc mesenteroides B-742CB. The 6.1-kb PstI fragments were ligated with pGEM-3Zf(-) and transformed into E. coli $DH5{\alpha}$. The recombinant clone (pDSRB742) synthesized dextran on an agar plate containing 2% (w/v) sucrose. The dextran synthesized was hydrolyzed with Penicillium endo-dextranase. The hydrolyzate was composed of glucose, isomaltose, isomaltotriose, and branced pentasaccharide. The nucleotide sequence of dsrB742 showed one open reading frame (ORF) composed of 4,524 bp encoding dextrasnsucrase. The deduced amino acid sequence revealed a calculated molecular mass of 168.6 kDa. It also showed an activity band of 184 kKa on a non-denaturing SDS-PAGE (10%). The amino acid sequence of DSRB742 exhibited a 50% similarity with DSRA from L. mesenteroides B-1299, a 70% similarity with DSRS from L. mesenteroides B-512 (F, FMCM) and a 45-56% similarity with Streptococcal GTFs.

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References

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