Transforming wheat vacuolar invertase into a high affinity sucrose:sucrose 1-fructosyltransferase

The New Phytologist
Lindsey SchroevenWim Van den Ende

Abstract

Vacuolar invertases (VIs) degrade sucrose to glucose and fructose. Additionally, the fructan plant wheat (Triticum aestivum) contains different fructosyltransferases (FTs), which have evolved from VIs by developing the capacity to bind sucrose or fructans as acceptor substrates. Modelling studies revealed a hydrogen bonding network in the conserved WMNDPNG motif of VIs, which is absent in FTs. In this study, the hydrogen bonding network of wheat VI was disrupted by site-directed mutagenesis in the 23WMNDPNG29 motif. While the single mutants (W23Y, N25S) showed a moderate increase in 1-kestose production, a synergistic effect was observed for the double mutant (W23Y+N25S), showing a 17-fold increase in transfructosylation capacity, and becoming a real sucrose:sucrose 1-fructosyltransferase. Vacuolar invertases are fully saturable enzymes, contrary to FTs. This is the first report on the development of a fully saturable FT with respect to 1-kestose formation. The superior kinetics (K(m) approximately 43 mM) make the enzyme useful for biotechnological applications. The results indicate that changes in the WMNDPNG motif are necessary to develop transfructosylating capability. The shift towards smaller and/or more hydrophilic residu...Continue Reading

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Citations

Aug 7, 2012·Plant Molecular Biology·Bao-Lam HuynhDelphine Fleury
Jan 9, 2009·Journal of Experimental Botany·Willem LammensWim Van den Ende
Sep 4, 2009·Journal of Experimental Botany·Lindsey SchroevenWim Van den Ende
Jul 19, 2011·Applied and Environmental Microbiology·Álvaro LafrayaJulia Marín-Navarro
Sep 1, 2015·Frontiers in Plant Science·Emerik De SadeleerWim Van den Ende
Nov 26, 2016·Plant Biotechnology Journal·Anna StolzeChristian Schulze Gronover
Sep 20, 2017·Plant, Cell & Environment·Maxime VersluysWim Van den Ende
Sep 22, 2009·The FEBS Journal·Wim Van den EndeKatrien Le Roy
Aug 20, 2017·Carbohydrate Polymers·Joran VerspreetChristophe M Courtin

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