title: The hexose transporters at the plasma membrane and the tonoplast of transformed plant cells: kinetic characterization of two distinct carriers.
abstract: The plasma membrane hexose transporter and the tonoplast hexose transporter from heterotrophically grown transformed Nicotiana tabacum cells have been studied in vitro using membrane vesicles for trans-zero transport studies. In highly purified phase-partitioned outside-out plasma membrane vesicles (PMV) the hexose transporter showed an apparent Km value of 230 microM (substrate: 3-O-methyl-D-glucose (3-OMG); pHi 7.2/pHo 7.2), which was reduced to 120 microM when a pH gradient was imposed (pHo 5.7/pHi 7.2). However, the Vmax value was not affected indicating that no stable pH gradient was formed. Uptake experiments with 14C-labelled acetate supported this interpretation. Transport was insensitive to N-ethylmaleimide (NEM; up to 1 mM concentration) and p-chloromercuribenzene sulfonate (PCMBS; up to 500 microM), whereas the tonoplast hexose transporter (in mixed inside / out and outside / out vesicles) was inhibited by NEM in a substrate-protectable manner, and PCMBS was also inhibitory. Kinetically two components with apparent Km values of 6 and 20 mM could be distinguished for the tonoplast hexose transporter. Substrate specificities of both transporters were similar except for D-galactose and D-fructose. The results indicate structural differences between the tonoplast and plasma membrane hexose transporters in plants.
authors.affiliation: Botanisches Institut, Johann Wolfgang Goethe Universität, Frankfurt F.R.G.
authors.forename: R
authors.initials: R
authors.lastname: Verstappen
authors.forename: S
authors.initials: S
authors.lastname: Ranostaj
authors.forename: T
authors.initials: T
authors.lastname: Rausch
mesh_terms: D019325:3-O-Methylglucose; D002731:4-Chloromercuribenzenesulfonate / Q000494:pharmacology; D000085:Acetates / Q000378:metabolism; D002462:Cell Membrane / Q000378:metabolism* / Q000648:ultrastructure; D005033:Ethylmaleimide / Q000494:pharmacology; D005632:Fructose / Q000378:metabolism; D005690:Galactose / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008757:Methylglucosides / Q000378:metabolism; D008854:Microscopy, Electron; D009004:Monosaccharide Transport Proteins / Q000378:metabolism*; D010944:Plants / Q000378:metabolism* / Q000648:ultrastructure; D010947:Plants, Toxic; D012231:Rhizobium; D014026:Nicotiana
chemical_list: D000085:Acetates; D008757:Methylglucosides; D009004:Monosaccharide Transport Proteins; D019325:3-O-Methylglucose; D005632:Fructose; D002731:4-Chloromercuribenzenesulfonate; D005033:Ethylmaleimide; D005690:Galactose