title: An evaluation of detergents for NMR structural studies of membrane proteins.
abstract: Structural information on membrane proteins lags far behind that on soluble proteins, in large part due to difficulties producing homogeneous, stable, structurally relevant samples in a membrane-like environment. In this study 25 membrane mimetics were screened using 2D (1)H-(15)N heteronuclear single quantum correlation NMR experiments to establish sample homogeneity and predict fitness for structure determination. A single detergent, 1-palmitoyl-2-hydroxy-sn-glycero-3-[phospho-RAC-(1-glycerol)] (LPPG), yielded high quality NMR spectra with sample lifetimes greater than one month for the five proteins tested - R. sphaeroides LH1 alpha and beta subunits, E. coli and B. pseudofirmus OF4 ATP synthase c subunits, and S. aureus small multidrug resistance transporter - with 1, 2, or 4 membrane spanning alpha-helices, respectively. Site-specific spin labeling established interhelical distances in the drug transporter and genetically fused dimers of c subunits in LPPG consistent with in vivo distances. Optical spectroscopy showed that LH1 beta subunits form native-like complexes with bacteriochlorophyll a in LPPG. All the protein/micelle complexes were estimated to exceed 100 kDaltons by translational diffusion measurements. However, analysis of (15)N transverse, longitudinal and (15)N[(1)H] nuclear Overhauser effect relaxation measurements yielded overall rotational correlation times of 8 to 12 nsec, similar to a 15-20 kDalton protein tumbling isotropically in solution, and consistent with the high quality NMR data observed.
authors.affiliation: Biochemistry Department, Albert Einstein College of Medicine, 1300 Morris Park Ave, Bronx, NY 10461, U.S.A.
authors.forename: Ray D
authors.initials: RD
authors.lastname: Krueger-Koplin
authors.forename: Paul L
authors.initials: PL
authors.lastname: Sorgen
authors.forename: Suzanne T
authors.initials: ST
authors.lastname: Krueger-Koplin
authors.forename: Iván O
authors.initials: IO
authors.lastname: Rivera-Torres
authors.forename: Sean M
authors.initials: SM
authors.lastname: Cahill
authors.forename: David B
authors.initials: DB
authors.lastname: Hicks
authors.forename: Leo
authors.initials: L
authors.lastname: Grinius
authors.forename: Terry A
authors.initials: TA
authors.lastname: Krulwich
authors.forename: Mark E
authors.initials: ME
authors.lastname: Girvin
mesh_terms: D017920:Antiporters / Q000737:chemistry / Q000494:pharmacology; D003902:Detergents / Q000737:chemistry* / Q000494:pharmacology; D029968:Escherichia coli Proteins; D005989:Glycerides / Q000737:chemistry* / Q000494:pharmacology; D008565:Membrane Proteins / Q000737:chemistry* / Q000187:drug effects / Q000494:pharmacology; D008823:Micelles; D025261:Mitochondrial Proton-Translocating ATPases / Q000737:chemistry / Q000187:drug effects; D019906:Nuclear Magnetic Resonance, Biomolecular; D017510:Protein Folding; D006180:Proton-Translocating ATPases / Q000737:chemistry; D013113:Spin Labels
chemical_list: C492788:1-palmitoyl-2-hydroxy-sn-glycero-3-(phospho-RAC-(1-glycerol)); D017920:Antiporters; D003902:Detergents; D029968:Escherichia coli Proteins; D005989:Glycerides; D008565:Membrane Proteins; D008823:Micelles; D013113:Spin Labels; C075436:EmrE protein, E coli; C103607:mitochondrial ATPase subunit c; D025261:Mitochondrial Proton-Translocating ATPases; D006180:Proton-Translocating ATPases