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Rewiring yeast sugar transporter preference through modifying a conserved protein motif

Proceedings of the National Academy of Sciences - PNAS, 2014-01, Vol.111 (1), p.131-136 [Peer Reviewed Journal]

copyright © 1993—2008 National Academy of Sciences of the United States of America ;Copyright National Academy of Sciences Jan 7, 2014 ;ISSN: 0027-8424 ;EISSN: 1091-6490 ;DOI: 10.1073/pnas.1311970111 ;PMID: 24344268

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Title:
Rewiring yeast sugar transporter preference through modifying a conserved protein motif
Author: Young, Eric M. ; Tong, Alice ; Bui, Hang ; Spofford, Caitlin ; Alper, Hal S.
Subjects: Amino Acid Motifs ; Biochemistry ; Bioengineering ; Biofuels ; Biological Sciences ; Biological Transport ; Biomass ; Candida - metabolism ; Candida intermedia ; Carbon ; Cloning, Molecular ; Debaryomyces hansenii ; Escherichia coli - metabolism ; Fermentation ; Fungal Proteins - chemistry ; Gene Expression Regulation, Fungal ; Genotype & phenotype ; Geometric growth ; Glucose - chemistry ; Hexoses ; Membranes ; Monosaccharide Transport Proteins - chemistry ; Monosaccharides ; Monosaccharides - chemistry ; Mutagenesis ; Mutation ; Phenotype ; Phenotypes ; Porters ; Proteins ; Saccharomyces cerevisiae ; Saccharomyces cerevisiae - metabolism ; Sugars ; Time Factors ; Transcriptional regulatory elements ; Xylose - chemistry ; Yeast ; Yeasts
Is Part Of: Proceedings of the National Academy of Sciences - PNAS, 2014-01, Vol.111 (1), p.131-136
Description: Utilization of exogenous sugars found in lignocellulosic biomass hydrolysates, such as xylose, must be improved before yeast can serve as an efficient biofuel and biochemical production platform. In particular, the first step in this process, the molecular transport of xylose into the cell, can serve as a significant flux bottleneck and is highly inhibited by other sugars. Here we demonstrate that sugar transport preference and kinetics can be rewired through the programming of a sequence motif of the general form G-G/F-XXX-G found in the first transmembrane span. By evaluating 46 different heterologously expressed transporters, we find that this motif is conserved among functional transporters and highly enriched in transporters that confer growth on xylose. Through saturation mutagenesis and subsequent rational mutagenesis, four transporter mutants unable to confer growth on glucose but able to sustain growth on xylose were engineered. Specifically, Candida intermedia gxs1 Phe ³⁸Ile ³⁹Met ⁴⁰, Scheffersomyces stipitis rgt2 Phe ³⁸ and Met ⁴⁰, and Saccharomyces cerevisiae hxt7 Ile ³⁹Met ⁴⁰Met ³⁴⁰ all exhibit this phenotype. In these cases, primary hexose transporters were rewired into xylose transporters. These xylose transporters nevertheless remained inhibited by glucose. Furthermore, in the course of identifying this motif, novel wild-type transporters with superior monosaccharide growth profiles were discovered, namely S. stipitis RGT2 and Debaryomyces hansenii 2D01474 . These findings build toward the engineering of efficient pentose utilization in yeast and provide a blueprint for reprogramming transporter properties.
Publisher: United States: National Academy of Sciences
Language: English
Identifier: ISSN: 0027-8424
EISSN: 1091-6490
DOI: 10.1073/pnas.1311970111
PMID: 24344268
Source: GFMER Free Medical Journals
MEDLINE
PubMed Central

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Rewiring yeast sugar transporter preference through modifying a conserved protein motif

copyright © 1993—2008 National Academy of Sciences of the United States of America ;Copyright National Academy of Sciences Jan 7, 2014 ;ISSN: 0027-8424 ;EISSN: 1091-6490 ;DOI: 10.1073/pnas.1311970111 ;PMID: 24344268

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