title: Rhizobium leguminosarum HupE is a highly-specific diffusion facilitator for nickel uptake.
abstract: Bacteria require nickel transporters for the synthesis of Ni-containing metalloenzymes in natural, low nickel habitats. In this work we carry out functional and topological characterization of Rhizobium leguminosarum HupE, a nickel permease required for the provision of this element for [NiFe] hydrogenase synthesis. Expression studies in the Escherichia coli nikABCDE mutant strain HYD723 revealed that HupE is a medium-affinity permease (apparent Km 227 ± 21 nM; Vmax 49 ± 21 pmol Ni(2+) min(-1) mg(-1) bacterial dry weight) that functions as an energy-independent diffusion facilitator for the uptake of Ni(ii) ions. This Ni(2+) transport is not inhibited by similar cations such as Mn(2+), Zn(2+), or Co(2+), but is blocked by Cu(2+). Analysis of site-directed HupE mutants allowed the identification of several residues (H36, D42, H43, F69, E90, H130, and E133) that are essential for HupE-mediated Ni uptake in E. coli cells. By using translational fusions to reporter genes we demonstrated the presence of five transmembrane domains with a periplasmic N-terminal domain and a C-terminal domain buried in the lipid bilayer. The periplasmic N-terminal domain contributes to stability and functionality of the protein.
authors: Departamento de Biotecnología, Escuela Técnica Superior de Ingenieros Agrónomos and Centro de Biotecnología y Genómica de Plantas (C.B.G.P.), Universidad Politécnica de Madrid, Campus de Montegancedo, Carretera M40-km 38, 28223 Pozuelo de Alarcón, Madrid, Spain. jose.palacios@upm.es.
authors: Marta
authors: M
authors: Albareda
authors: Agnès
authors: A
authors: Rodrigue
authors: Belén
authors: B
authors: Brito
authors: Tomás
authors: T
authors: Ruiz-Argüeso
authors: Juan
authors: J
authors: Imperial
authors: Marie-Andrée
authors: MA
authors: Mandrand-Berthelot
authors: Jose
authors: J
authors: Palacios
mesh_terms: D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry*; D004926:Escherichia coli / Q000378:metabolism; D017930:Genes, Reporter; D006864:Hydrogenase / Q000737:chemistry*; D008024:Ligands; D008051:Lipid Bilayers / Q000737:chemistry; D008565:Membrane Proteins / Q000737:chemistry*; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D009532:Nickel / Q000737:chemistry*; D017434:Protein Structure, Tertiary; D016961:Rhizobium leguminosarum / Q000737:chemistry*
chemical_list: D001426:Bacterial Proteins; D008024:Ligands; D008051:Lipid Bilayers; D008565:Membrane Proteins; C074902:HupE protein, Rhizobium leguminosarum; D009532:Nickel; D006864:Hydrogenase