Enhanced levan production using chitin-binding domain fused levansucrase immobilized on chitin beads

Appl Microbiol Biotechnol. 2009 Mar;82(3):445-51. doi: 10.1007/s00253-008-1772-z. Epub 2008 Nov 19.

Abstract

Levan is a homopolymer of fructose which can be produced by the transfructosylation reaction of levansucrase (EC 2.4.1.10) from sucrose. In particular, levan synthesized by Zymomonas mobilis has found a wide and potential application in the food and pharmaceutical industry. In this study, the immobilization of Z. mobilis levansucrae (encoded by levU) was attempted for repeated production of levan. By fusion levU with the chitin-binding domain (ChBD), the hybrid protein was overproduced in a soluble form in Escherichia coli. After direct absorption of the protein mixture from E. coli onto chitin beads, levansucrase tagged with ChBD was found to specifically attach to the affinity matrix. Subsequent analysis indicated that the linkage between the enzyme and chitin beads was substantially stable. Furthermore, with 20% sucrose, the production of levan was enhanced by 60% to reach 83 g/l using the immobilized levansucrase as compared to that by the free counterpart. This production yield accounts for 41.5% conversion yield (g/g) on the basis of sucrose. After all, a total production of levan with 480 g/l was obtained by recycling of the immobilized enzyme for seven times. It is apparent that this approach offers a promising way for levan production by Z. mobilis levansucrase immobilized on chitin beads.

Publication types

  • Evaluation Study
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Biomass
  • Chitin / chemistry*
  • Enzyme Stability
  • Enzymes, Immobilized / chemistry
  • Enzymes, Immobilized / genetics
  • Enzymes, Immobilized / isolation & purification
  • Enzymes, Immobilized / metabolism*
  • Escherichia coli / genetics
  • Escherichia coli / metabolism
  • Fructans / metabolism*
  • Hexosyltransferases / chemistry
  • Hexosyltransferases / genetics
  • Hexosyltransferases / isolation & purification
  • Hexosyltransferases / metabolism*
  • Protein Engineering / methods*
  • Protein Structure, Tertiary
  • Recombinant Fusion Proteins / chemistry
  • Recombinant Fusion Proteins / genetics
  • Recombinant Fusion Proteins / isolation & purification
  • Recombinant Fusion Proteins / metabolism
  • Zymomonas / enzymology

Substances

  • Enzymes, Immobilized
  • Fructans
  • Recombinant Fusion Proteins
  • Chitin
  • levan
  • Hexosyltransferases
  • levansucrase