Expression and activity analysis of sucrose:sucrose 1-fructosyltransferase from onion

N Biotechnol. 2010 Sep 30;27(4):324-9. doi: 10.1016/j.nbt.2010.02.004. Epub 2010 Feb 25.

Abstract

This study was designed to express the onion fructosyltransferase by Escherichia coli DH5alpha, and obtain the optimal conditions of FST-1 activity. Thereby, fructosyltransferase gene was obtained by RT-PCR from onion in this experiment, and named FST-1. The expressed proteins were analyzed by SDS-PAGE. FST-1 activity was identified by the high performance liquid chromatography (HPLC). The optimal conditions of FST-1 were analyzed by the dinonylnaphthalene sulfonic acid (DNS) and orthogonal test. Results revealed that FST-1 was identified to 98% similarity with fructosyltransferase mRNA of onion (accession number: AJ006066). FST-1 was successfully expressed in E. coli DH5alpha. HPLC results indicated that the expressed protein from FST-1 had a good transferring activity for fructose. The optimal conditions of FST-1 in catalyzing reaction were the pH 5.0, 45 degrees C and 60% sucrose substrate. The results in this experiment would lay the foundation for the large-scale of kestose by bio-catalysis method.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Chromatography, High Pressure Liquid
  • Gene Expression Regulation, Enzymologic
  • Gene Expression Regulation, Plant
  • Hexosyltransferases / genetics
  • Hexosyltransferases / metabolism*
  • Hydrogen-Ion Concentration
  • Onions / enzymology*
  • Onions / genetics
  • Reference Standards
  • Reverse Transcriptase Polymerase Chain Reaction
  • Substrate Specificity
  • Temperature
  • Time Factors

Substances

  • Hexosyltransferases
  • inulosucrase