title: Probing Structural Dynamics and Topology of the KCNE1 Membrane Protein in Lipid Bilayers via Site-Directed Spin Labeling and Electron Paramagnetic Resonance Spectroscopy.
abstract: KCNE1 is a single transmembrane protein that modulates the function of voltage-gated potassium channels, including KCNQ1. Hereditary mutations in the genes encoding either protein can result in diseases such as congenital deafness, long QT syndrome, ventricular tachyarrhythmia, syncope, and sudden cardiac death. Despite the biological significance of KCNE1, the structure and dynamic properties of its physiologically relevant native membrane-bound state are not fully understood. In this study, the structural dynamics and topology of KCNE1 in bilayered lipid vesicles was investigated using site-directed spin labeling (SDSL) and electron paramagnetic resonance (EPR) spectroscopy. A 53-residue nitroxide EPR scan of the KCNE1 protein sequence including all 27 residues of the transmembrane domain (45-71) and 26 residues of the N- and C-termini of KCNE1 in lipid bilayered vesicles was analyzed in terms of nitroxide side-chain motion. Continuous wave-EPR spectral line shape analysis indicated the nitroxide spin label side-chains located in the KCNE1 TMD are less mobile when compared to the extracellular region of KCNE1. The EPR data also revealed that the C-terminus of KCNE1 is more mobile when compared to the N-terminus. EPR power saturation experiments were performed on 41 sites including 18 residues previously proposed to reside in the transmembrane domain (TMD) and 23 residues of the N- and C-termini to determine the topology of KCNE1 with respect to the 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC)/1-palmitoyl-2-oleoyl-sn-glycero-3-phospho-(1'-rac-glycerol) (POPG) lipid bilayers. The results indicated that the transmembrane domain is indeed buried within the membrane, spanning the width of the lipid bilayer. Power saturation data also revealed that the extracellular region of KCNE1 is solvent-exposed with some of the portions partially or weakly interacting with the membrane surface. These results are consistent with the previously published solution NMR structure of KCNE1 in micelles.
authors.affiliation: Department of Chemistry and Biochemistry, Miami University , Oxford, Ohio 45056, United States.
authors.forename: Indra D
authors.initials: ID
authors.lastname: Sahu
authors.affiliation: Department of Chemistry and Biochemistry, Miami University , Oxford, Ohio 45056, United States.
authors.forename: Andrew F
authors.initials: AF
authors.lastname: Craig
authors.affiliation: Department of Chemistry and Biochemistry, Miami University , Oxford, Ohio 45056, United States.
authors.forename: Megan M
authors.initials: MM
authors.lastname: Dunagan
authors.affiliation: Department of Chemistry and Biochemistry, Miami University , Oxford, Ohio 45056, United States.
authors.forename: Kaylee R
authors.initials: KR
authors.lastname: Troxel
authors.affiliation: Department of Chemistry and Biochemistry, Miami University , Oxford, Ohio 45056, United States.
authors.forename: Rongfu
authors.initials: R
authors.lastname: Zhang
authors.affiliation: Department of Chemistry and Biochemistry, Miami University , Oxford, Ohio 45056, United States.
authors.forename: Andrew G
authors.initials: AG
authors.lastname: Meiberg
authors.affiliation: Department of Chemistry and Biochemistry, Miami University , Oxford, Ohio 45056, United States.
authors.forename: Corrinne N
authors.initials: CN
authors.lastname: Harmon
authors.affiliation: Department of Chemistry and Biochemistry, Miami University , Oxford, Ohio 45056, United States.
authors.forename: Robert M
authors.initials: RM
authors.lastname: McCarrick
authors.affiliation: Department of Biochemistry and Center for Structural Biology, Vanderbilt University , Nashville, Tennessee 37232, United States.
authors.forename: Brett M
authors.initials: BM
authors.lastname: Kroncke
authors.affiliation: Department of Biochemistry and Center for Structural Biology, Vanderbilt University , Nashville, Tennessee 37232, United States.
authors.forename: Charles R
authors.initials: CR
authors.lastname: Sanders
authors.affiliation: Department of Chemistry and Biochemistry, Miami University , Oxford, Ohio 45056, United States.
authors.forename: Gary A
authors.initials: GA
authors.lastname: Lorigan
mesh_terms: D000595:Amino Acid Sequence; D004578:Electron Spin Resonance Spectroscopy; D008051:Lipid Bilayers / Q000737:chemistry*; D056004:Molecular Dynamics Simulation; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D024642:Potassium Channels, Voltage-Gated / Q000737:chemistry* / Q000235:genetics; D011487:Protein Conformation
chemical_list: D008051:Lipid Bilayers; D024642:Potassium Channels, Voltage-Gated