title: Solution-NMR characterization of outer-membrane protein A from E. coli in lipid bilayer nanodiscs and detergent micelles.
abstract: X-ray crystallography and solution NMR of detergent-reconstituted OmpA (outer membrane protein A from E. coli) had shown that this protein forms an eight-stranded transmembrane β-barrel, but only limited information was obtained for the extracellular loops. In NMR studies of OmpA in two different detergent micelles, "NMR-invisible" amino acid residues in-between the extracellular loops and the β-barrel prevented complete structural characterization. Here, we show that this NMR-invisible ring around the β-barrel of OmpA is also present in lipid bilayer nanodiscs and in mixed micelles with a third detergent, thus suggesting that the implicated rate processes have a functional role rather than representing an artifact of the protein reconstitution. In addition to sequence-specific NMR assignments for OmpA in the nanodiscs, the present results are based on a protocol of micro-coil TROSY- and CRINEPT-type NMR diffusion measurements for studying the hydrodynamic properties and the foldedness of [(2)H,(15)N]-labeled membrane proteins in nanodiscs. This protocol can be applied under conditions closely similar to those used for NMR structure determinations or crystallization trials.
authors.affiliation: Department of Integrative Structural and Computational Biology, The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, CA 92037 (USA).
authors.forename: Lukas
authors.initials: L
authors.lastname: Sušac
authors.forename: Reto
authors.initials: R
authors.lastname: Horst
authors.forename: Kurt
authors.initials: K
authors.lastname: Wüthrich
mesh_terms: D001425:Bacterial Outer Membrane Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D003902:Detergents / Q000737:chemistry; D004926:Escherichia coli / Q000378:metabolism*; D008051:Lipid Bilayers / Q000737:chemistry* / Q000378:metabolism; D009682:Magnetic Resonance Spectroscopy; D008823:Micelles; D049329:Nanostructures / Q000737:chemistry*; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics; D012996:Solutions / Q000737:chemistry
chemical_list: D001425:Bacterial Outer Membrane Proteins; D003902:Detergents; D008051:Lipid Bilayers; D008823:Micelles; D011993:Recombinant Fusion Proteins; D012996:Solutions; C078167:OMPA outer membrane proteins
keywords: NMR spectroscopy; membrane protein reconstitution; membrane proteins; nanodiscs; structural biology