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A urea channel from Bacillus cereus reveals a novel hexameric structure

DOI: 10.1042/BJ20120169 DOI Help

Authors: Gerard H. M. Huysmans (University of Leeds) , Nathan Chan (University of Sheffield) , Jocelyn M. Baldwin (University of Leeds) , Vincent L. G. Postis (Leeds University) , Svetomir B. Tzokov (University of Sheffield) , Sarah E. Deacon (University of Leeds) , Sylvia Y. M. Yao (University of Alberta) , James D. Young (University of Alberta) , Michael J. Mcpherson (University of Leeds) , Per A. Bullough (University of Sheffield) , Stephen A. Baldwin (University of Leeds)
Co-authored by industrial partner: No

Type: Journal Paper
Journal: Biochemical Journal , VOL 445 (2)

State: Published (Approved)
Published: July 2012

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 ?-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.

Journal Keywords: Cryo-Electron Microscopy; Electron Crystallography; Helicobacter Pylori; Membrane; Protein Structure; Urea Channel

Subject Areas: Biology and Bio-materials

Diamond Offline Facilities: Membrane Protein Laboratory (MPL)

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