title: Defining membrane spanning domains and crucial membrane-localized acidic amino acid residues for K⁺ transport of a Kup/HAK/KT-type Escherichia coli potassium transporter.
abstract: Potassium (K(+))-uptake transport proteins present in prokaryote and eukaryote cells are categorized into two classes; Trk/Ktr/HKT, K(+) channel, and Kdp belong to the same superfamily, whereas the remaining K(+)-uptake family, Kup/HAK/KT has no homology to the others, and neither its membrane topology nor crucial residues for K(+) uptake have been identified. We examined the topology of Kup from Escherichia coli. Results from the reporter fusion and cysteine labeling assays support a model with 12 membrane-spanning domains. A model for proton-coupled K(+) uptake mediated by Kup has been proposed. However, this study did not show any stimulation of Kup activity at low pH and any evidence of involvement of the three His in Kup-mediated K(+) uptake. Moreover, replacement of all four cysteines of Kup with serine did not abolish K(+) transport activity. To gain insight on crucial residues of Kup-mediated K(+) uptake activity, we focused on acidic residues in the predicted external and transmembrane regions, and identified four residues in the membrane regions required for K(+) uptake activity. This is different from no membrane-localized acidic residues essential for Trk/Ktr/HKTs, K(+) channels and Kdp. Taken together, these results demonstrate that Kup belongs to a distinct type of K(+) transport system.
authors: Department of Biomolecular Engineering, Graduate School of Engineering, Tohoku University, Aobayama 6-6-07, Sendai 980-8579, Japan; Department of Chemistry and Biotechnology, Graduate School of Natural Science and Technology, Okayama University, Okayama 700-8530, Japan; and Division of Biological Sciences, Cell and Developmental Biology Section, University of California at San Diego, La Jolla, CA 92093-0116, USA.
authors: Yoko
authors: Y
authors: Sato
authors: Kei
authors: K
authors: Nanatani
authors: Shin
authors: S
authors: Hamamoto
authors: Makoto
authors: M
authors: Shimizu
authors: Miho
authors: M
authors: Takahashi
authors: Mayumi
authors: M
authors: Tabuchi-Kobayashi
authors: Akifumi
authors: A
authors: Mizutani
authors: Julian I
authors: JI
authors: Schroeder
authors: Satoshi
authors: S
authors: Souma
authors: Nobuyuki
authors: N
authors: Uozumi
mesh_terms: D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D001692:Biological Transport; D002462:Cell Membrane / Q000378:metabolism*; D003545:Cysteine / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics / Q000254:growth & development / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D006639:Histidine / Q000235:genetics; D006863:Hydrogen-Ion Concentration; D008969:Molecular Sequence Data; D011188:Potassium / Q000378:metabolism; D017434:Protein Structure, Tertiary; D012694:Serine / Q000235:genetics
chemical_list: D029968:Escherichia coli Proteins; C483828:kup protein, E coli; D012694:Serine; D006639:Histidine; D003545:Cysteine; D011188:Potassium
keywords: Escherichia coli; Kup; acidic amino acid residues; potassium; topology