Structural analysis of bioengineered alpha-D-glucan produced by a triple mutant of the Glucansucrase GTF180 enzyme from Lactobacillus reuteri strain 180: generation of (alpha1-->4) linkages in a native (1-->3)(1-->6)-alpha-D-glucan

Biomacromolecules. 2008 Aug;9(8):2251-8. doi: 10.1021/bm800410w. Epub 2008 Jul 11.

Abstract

Site-directed mutagenesis of the glucansucrase gtf180 gene from Lactobacillus reuteri strain 180 was used to transform the active site region. The alpha-D-glucan ( mEPS-PNNS) produced by the triple mutant V1027P:S1137N:A1139S differed in structure from that of the wild-type alpha-D-glucan ( EPS180). Besides (alpha1-->3) and (alpha1-->6) linkages, as present in EPS180, mEPS-PNNS also contained (alpha1-->4) linkages. Linkage analysis, periodate oxidation, and 1D/2D (1)H NMR spectroscopy of the intact mEPS-PNNS, as well as MS and NMR analysis of oligosaccharides obtained by partial acid hydrolysis of mEPS-PNNS afforded a composite model, which includes all identified structural features.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Anions
  • Chromatography, Ion Exchange / methods
  • Glucans
  • Glycosyltransferases / chemistry*
  • Glycosyltransferases / genetics*
  • Hydrogen-Ion Concentration
  • Hydrolysis
  • Lactic Acid / metabolism
  • Limosilactobacillus reuteri / enzymology*
  • Limosilactobacillus reuteri / genetics*
  • Magnetic Resonance Spectroscopy
  • Mass Spectrometry / methods
  • Mutagenesis, Site-Directed
  • Mutation*
  • Oxygen / chemistry
  • Periodic Acid / chemistry
  • Protein Engineering / methods

Substances

  • Anions
  • Glucans
  • Periodic Acid
  • Lactic Acid
  • metaperiodate
  • Glycosyltransferases
  • alternansucrase
  • Oxygen