title: Novel transporters from Kluyveromyces marxianus and Pichia guilliermondii expressed in Saccharomyces cerevisiae enable growth on L-arabinose and D-xylose.
abstract: Genes encoding L-arabinose transporters in Kluyveromyces marxianus and Pichia guilliermondii were identified by functional complementation of Saccharomyces cerevisiae whose growth on L-arabinose was dependent on a functioning L-arabinose transporter, or by screening a differential display library, respectively. These transporters also transport D-xylose and were designated KmAXT1 (arabinose-xylose transporter) and PgAXT1, respectively. Transport assays using L-arabinose showed that KmAxt1p has K(m) 263 mM and V(max) 57 nM/mg/min, and PgAxt1p has K(m) 0.13 mM and V(max) 18 nM/mg/min. Glucose, galactose and xylose significantly inhibit L-arabinose transport by both transporters. Transport assays using D-xylose showed that KmAxt1p has K(m) 27 mM and V(max) 3.8 nM/mg/min, and PgAxt1p has K(m) 65 mM and V(max) 8.7 nM/mg/min. Neither transporter is capable of recovering growth on glucose or galactose in a S. cerevisiae strain deleted for hexose and galactose transporters. Transport kinetics of S. cerevisiae Gal2p showed K(m) 371 mM and V(max) 341 nM/mg/min for L-arabinose, and K(m) 25 mM and V(max) 76 nM/mg/min for galactose. Due to the ability of Gal2p and these two newly characterized transporters to transport both L-arabinose and D-xylose, one scenario for the complete usage of biomass-derived pentose sugars would require only the low-affinity, high-throughput transporter Gal2p and one additional high-affinity general pentose transporter, rather than dedicated D-xylose or L-arabinose transporters. Additionally, alignment of these transporters with other characterized pentose transporters provides potential targets for substrate recognition engineering.
authors.affiliation: National Renewable Energy Laboratory, National Bioenergy Centre, Golden, CO, USA.
authors.forename: Eric P
authors.initials: EP
authors.lastname: Knoshaug
authors.affiliation: VTT Technical Research Centre of Finland, PO Box 1000, FI-02044 VTT, Finland.
authors.forename: Virve
authors.initials: V
authors.lastname: Vidgren
authors.affiliation: VTT Technical Research Centre of Finland, PO Box 1000, FI-02044 VTT, Finland.
authors.forename: Frederico
authors.initials: F
authors.lastname: Magalhães
authors.affiliation: National Renewable Energy Laboratory, National Bioenergy Centre, Golden, CO, USA.
authors.forename: Eric E
authors.initials: EE
authors.lastname: Jarvis
authors.affiliation: National Renewable Energy Laboratory, National Bioenergy Centre, Golden, CO, USA.
authors.forename: Mary Ann
authors.initials: MA
authors.lastname: Franden
authors.affiliation: National Renewable Energy Laboratory, National Bioenergy Centre, Golden, CO, USA.
authors.forename: Min
authors.initials: M
authors.lastname: Zhang
authors.affiliation: National Renewable Energy Laboratory, National Bioenergy Centre, Golden, CO, USA.
authors.forename: Arjun
authors.initials: A
authors.lastname: Singh
mesh_terms: D001089:Arabinose / Q000737:chemistry / Q000378:metabolism*; D001692:Biological Transport; D005656:Fungal Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D015870:Gene Expression; D005818:Genetic Engineering; D007700:Kinetics; D007716:Kluyveromyces / Q000235:genetics*; D026901:Membrane Transport Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D010843:Pichia / Q000235:genetics*; D012441:Saccharomyces cerevisiae / Q000737:chemistry / Q000235:genetics / Q000254:growth & development* / Q000378:metabolism; D014994:Xylose / Q000737:chemistry / Q000378:metabolism*
chemical_list: D005656:Fungal Proteins; D026901:Membrane Transport Proteins; D014994:Xylose; D001089:Arabinose
keywords: GAL2; arabinose; transport; xylose; yeast