ZrFsy1, a high-affinity fructose/H+ symporter from fructophilic yeast Zygosaccharomyces rouxii

PLoS One. 2013 Jul 2;8(7):e68165. doi: 10.1371/journal.pone.0068165. Print 2013.

Abstract

Zygosaccharomyces rouxii is a fructophilic yeast than can grow at very high sugar concentrations. We have identified an ORF encoding a putative fructose/H(+) symporter in the Z. rouxii CBS 732 genome database. Heterologous expression of this ORF in a S. cerevisiae strain lacking its own hexose transporters (hxt-null) and subsequent kinetic characterization of its sugar transport activity showed it is a high-affinity low-capacity fructose/H(+) symporter, with Km 0.45 ± 0.07 mM and Vmax 0.57 ± 0.02 mmol h(-1) (gdw)(-1). We named it ZrFsy1. This protein also weakly transports xylitol and sorbose, but not glucose or other hexoses. The expression of ZrFSY1 in Z. rouxii is higher when the cells are cultivated at extremely low fructose concentrations (<0.2%) and on non-fermentable carbon sources such as mannitol and xylitol, where the cells have a prolonged lag phase, longer duplication times and change their microscopic morphology. A clear phenotype was determined for the first time for the deletion of a fructose/H(+) symporter in the genome where it occurs naturally. The effect of the deletion of ZrFSY1 in Z. rouxii cells is only evident when the cells are cultivated at very low fructose concentrations, when the ZrFsy1 fructose symporter is the main active fructose transporter system.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Biological Transport / genetics
  • Fructose / metabolism*
  • Fungal Proteins / genetics
  • Fungal Proteins / metabolism*
  • Gene Deletion
  • Gene Expression Regulation, Fungal
  • Green Fluorescent Proteins / genetics
  • Green Fluorescent Proteins / metabolism
  • Microscopy, Fluorescence
  • Molecular Sequence Data
  • Monosaccharide Transport Proteins / genetics
  • Monosaccharide Transport Proteins / metabolism*
  • Phylogeny
  • Protons
  • Reverse Transcriptase Polymerase Chain Reaction
  • Saccharomyces cerevisiae / genetics
  • Saccharomyces cerevisiae / metabolism
  • Sequence Homology, Amino Acid
  • Symporters / classification
  • Symporters / genetics
  • Symporters / metabolism*
  • Zygosaccharomyces / genetics
  • Zygosaccharomyces / metabolism*

Substances

  • Fungal Proteins
  • Monosaccharide Transport Proteins
  • Protons
  • Symporters
  • Green Fluorescent Proteins
  • Fructose

Associated data

  • GENBANK/AAA34701
  • GENBANK/AAB64778
  • GENBANK/AAB68933
  • GENBANK/AAU87358
  • GENBANK/CAC08232
  • GENBANK/CAC79614
  • GENBANK/CAD56485
  • GENBANK/CAR26745
  • GENBANK/CAR28354
  • GENBANK/CAR31108
  • GENBANK/CAY86682
  • GENBANK/CCI61473
  • GENBANK/CCI61478
  • GENBANK/CCI61480
  • GENBANK/DAA06788
  • GENBANK/DAA06790
  • GENBANK/DAA10243
  • GENBANK/DAA12185
  • RefSeq/XP_001794684
  • RefSeq/XP_001935732
  • RefSeq/XP_389511
  • RefSeq/XP_453656

Grant support

This work was supported by Fundação para a Ciência e Tecnologia, Portugal (Post-Doctoral fellowships SFRH/BPD/41812/2007 to M.J.L. and SFRH/BPD/85143/2012 to C.P., FCT Ciência 2007 and Research Project ”Improvement of fructose fermentation by industrial Saccharomyces cerevisiae strains„ (PTDC/AGR-ALI/112802/2009)) and Czech MSMT LC531, GACR P503/10/0307, AV0Z 50110509 and RVO 6798582 grants. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.