title: Topology and immersion depth of an integral membrane protein by paramagnetic rates from dissolved oxygen.
abstract: In studies of membrane proteins, knowledge of protein topology can provide useful insight into both structure and function. In this work, we present a solution NMR method for the measurement the tilt angle and average immersion depth of alpha helices in membrane proteins, from analysis of the paramagnetic relaxation rate enhancements arising from dissolved oxygen. No modification to the micelle or protein is necessary, and the topology of both transmembrane and amphipathic helices are readily determined. We apply this method to the measure the topology of a monomeric mutant of phospholamban (AFA-PLN), a 52-residue membrane protein containing both an amphipathic and a transmembrane alpha helix. In dodecylphosphocholine micelles, the amphipathic helix of AFA-PLN was found to have a tilt angle of 87° ± 1° and an average immersion depth of 13.2 Å. The transmembrane helix was found to have an average immersion depth of 5.4 Å, indicating residues 41 and 42 are closest to the micelle centre. The resolution of paramagnetic relaxation rate enhancements from dissolved oxygen compares favourably to those from Ni (II), a hydrophilic paramagnetic species.
authors.affiliation: Department of Chemistry, University of Toronto Mississauga, Mississauga, ON L5L 1C6, Canada.
authors.forename: M Sameer
authors.initials: MS
authors.lastname: Al-Abdul-Wahid
authors.forename: Raffaello
authors.initials: R
authors.lastname: Verardi
authors.forename: Gianluigi
authors.initials: G
authors.lastname: Veglia
authors.forename: R Scott
authors.initials: RS
authors.lastname: Prosser
mesh_terms: D002135:Calcium-Binding Proteins / Q000737:chemistry / Q000378:metabolism; D057927:Hydrophobic and Hydrophilic Interactions; D007700:Kinetics; D008565:Membrane Proteins / Q000737:chemistry*; D008823:Micelles; D019906:Nuclear Magnetic Resonance, Biomolecular / Q000379:methods*; D010100:Oxygen / Q000737:chemistry* / Q000378:metabolism; D010767:Phosphorylcholine / Q000031:analogs & derivatives; D012996:Solutions; D000099211:Phospholamban
chemical_list: D002135:Calcium-Binding Proteins; D008565:Membrane Proteins; D008823:Micelles; D010100:Oxygen; D010767:Phosphorylcholine; D012996:Solutions; D000099211:Phospholamban; C028810:dodecylphosphocholine