title: A urea channel from Bacillus cereus reveals a novel hexameric structure.
abstract: Urea is exploited as a nitrogen source by bacteria, and its breakdown products, ammonia and bicarbonate, are employed to counteract stomach acidity in pathogens such as Helicobacter pylori. Uptake in the latter is mediated by UreI, a UAC (urea amide channel) family member. In the present paper, we describe the structure and function of UACBc, a homologue from Bacillus cereus. The purified channel was found to be permeable not only to urea, but also to other small amides. CD and IR spectroscopy revealed a structure comprising mainly α-helices, oriented approximately perpendicular to the membrane. Consistent with this finding, site-directed fluorescent labelling indicated the presence of seven TM (transmembrane) helices, with a cytoplasmic C-terminus. In detergent, UACBc exists largely as a hexamer, as demonstrated by both cross-linking and size-exclusion chromatography. A 9 Å (1 Å=0.1 nm) resolution projection map obtained by cryo-electron microscopy of two-dimensional crystals shows that the six protomers are arranged in a planar hexameric ring. Each exhibits six density features attributable to TM helices, surrounding a putative central channel, while an additional helix is peripherally located. Bioinformatic analyses allowed individual TM regions to be tentatively assigned to the density features, with the resultant model enabling identification of residues likely to contribute to channel function.
authors.affiliation: Astbury Centre for Structural Molecular Biology, Institute of Membrane and Systems Biology, University of Leeds, Leeds LS2 9JT, U.K.
authors.forename: Gerard H M
authors.initials: GH
authors.lastname: Huysmans
authors.forename: Nathan
authors.initials: N
authors.lastname: Chan
authors.forename: Jocelyn M
authors.initials: JM
authors.lastname: Baldwin
authors.forename: Vincent L G
authors.initials: VL
authors.lastname: Postis
authors.forename: Svetomir B
authors.initials: SB
authors.lastname: Tzokov
authors.forename: Sarah E
authors.initials: SE
authors.lastname: Deacon
authors.forename: Sylvia Y M
authors.initials: SY
authors.lastname: Yao
authors.forename: James D
authors.initials: JD
authors.lastname: Young
authors.forename: Michael J
authors.initials: MJ
authors.lastname: McPherson
authors.forename: Per A
authors.initials: PA
authors.lastname: Bullough
authors.forename: Stephen A
authors.initials: SA
authors.lastname: Baldwin
mesh_terms: D000595:Amino Acid Sequence; D001409:Bacillus cereus / Q000378:metabolism*; D001426:Bacterial Proteins / Q000737:chemistry* / Q000378:metabolism; D020285:Cryoelectron Microscopy; D007473:Ion Channels / Q000737:chemistry* / Q000378:metabolism; D026901:Membrane Transport Proteins / Q000737:chemistry* / Q000378:metabolism; D008969:Molecular Sequence Data; D017433:Protein Structure, Secondary; D017386:Sequence Homology, Amino Acid; D014508:Urea / Q000737:chemistry / Q000378:metabolism*
chemical_list: D001426:Bacterial Proteins; D007473:Ion Channels; D026901:Membrane Transport Proteins; D014508:Urea