Abstract
NMR is an essential tool to characterize the structure, dynamics, and interactions of biomolecules at an atomic level. Its application to membrane protein (MP) structure determination is challenging and currently an active and rapidly developing field. Main difficulties are the low sensitivity of the technique, the size limitation, and the intrinsic motional properties of the system under investigation. Solution and solid-state NMR (ssNMR) have common and own specific requirements. Solution NMR requires a careful choice of the detergent, elaborated stable isotope labelling schemes to overcome signal overlaps and to collect distance restraints. Excessive spectra crowding hampered large MP structure determination by ssNMR, and so far only high resolution structure of small or fragments of MP have been determined. However, ssNMR provides the unique opportunity to obtain atomic level information of MP in phospholipid bilayers such as orientation of the protein in the membrane. Specific and careful sample preparations are required in combination with uniformly and partially labelled protein for ssNMR spectra assignment. Distance restraints measurements benefit from methodologies currently developed for small soluble proteins in micro-crystalline state.
Recent advances in the field increased the releasing rate of high resolution MP structures, providing unprecedented structural and dynamics information making NMR a powerful tool for structural and functional membrane protein studies.
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References
Vold RR, Prosser RS (1996) Magnetically oriented phospholipid bilayered micelles for structural studies of polypeptides. Does the ideal bicelle exist? J Magn Res B 113:267–271
Tugarinov V, Hwang PM, Kay LE (2004) Nuclear magnetic resonance spectroscopy of high-molecular-weight proteins. Annu Rev Biochem 73:107–146
Page RC, Li C, Hu J, Gao FP, Cross TA (2007) Lipid bilayers: an essential environment for the understanding of membrane proteins. Magn Reson Chem 45:S2–S11
De Angelis AA, Jones DH, Grant CV, Park SH, Mesleh MF, Opella SJ (2005) NMR experiments on aligned samples of membrane proteins. Methods Enzymol 394:350–382
Watts A, Straus SK, Grage SL, Kamihira M, Lam YH, Zhao X (2004) Membrane protein structure determination using solid-state NMR Methods. Mol Biol 278:403–473
Nielsen N, Malmendal A, Vosegaard T (2004) Techniques and applications of NMR to membrane proteins. Mol Membr Biol 21:129–141
Gordon E, Horsefield R, Swarts HG, de Pont JJ, Neutze R, Snijder A (2008) Effective high-throughput overproduction of membrane proteins in Escherichia coli. Protein Expr Purif 62:1–8
Tyler RC, Sreenath HK, Singh S, Aceti DJ, Bingman CA, Markley JL, Fox BG (2005) Auto-induction medium for the production of [U-15N]- and [U-13C, U-15N]-labeled proteins for NMR screening and structure determination. Protein Expr Purif 40:268–278
Koglin A, Klammt C, Trbovic N, Schwarz D, Schneider B, Schafer B, Lohr F, Bernhard F, Dotsch V (2006) Combination of cell-free expression and NMR spectroscopy as a new approach for structural investigation of membrane proteins. Magn Reson Chem 44:S17–S23
Reckel S, Sobhanifar S, Schneider B, Junge F, Schwarz D, Durst F, Lohr F, Guntert P, Bernhard F, Dotsch V (2008) Transmembrane segment enhanced labeling as a tool for the back bone assignment of alpha-helical membrane proteins. Proc Natl Acad Sci U S A 105:8262–8267
Krueger-Koplin RD, Sorgen PL, Krueger-Koplin ST, Rivera-Torres IO, Cahill SM, Hicks DB, Grinius L, Krulwich TA, Girvin ME (2004) An evaluation of detergents for NMR structural studies of membrane proteins. J Biomol NMR 28:43–57
Page RC, Moore JD, Nguyen HB, Sharma M, Chase R, Gao FP, Mobley CK, Sanders CR, Ma L, Sonnichsen FD, Lee S, Howell SC, Opella SJ, Cross TA (2006) Comprehensive evaluation of solution nuclear magnetic resonance spectroscopy sample preparation for helical integral membrane proteins. J Struct Funct Genomics 7:51–64
Zhang Q, Horst R, Geralt M, Ma X, Hong WX, Finn MG, Stevens RC, Wuthrich K (2008) Microscale NMR screening of new detergents for membrane protein structural biology. J Am Chem Soc 130:7357–7363
Sanders CR, Sonnichsen F (2006) Solution NMR of membrane proteins: practice and challenges. Magn Reson Chem 44:S24–S40
Poget SF, Girvin ME (2007) Solution NMR of membrane proteins in bilayer mimics: small is beautiful, but sometimes bigger is better. Biochim Biophys Acta 1768:3098–3106
Bocharov EV, Pustovalova YE, Pavlov KV, Volynsky PE, Goncharuk MV, Ermolyuk YS, Karpunin DV, Schulga AA, Kirpichnikov MP, Efremov RG, Maslennikov IV, Arseniev AS (2007) Unique dimeric structure of BNip3 transmembrane domain suggests membrane permeabilization as a cell death trigger. J Biol Chem 282:16256–16266
Shih SC, Stoica I, Goto NK (2008) Investigation of the utility of selective methyl protonation for determination of membrane protein structures. J Biomol NMR 42:49–58
Zhou Y, Cierpicki T, Flores Jimenez RH, Lukasik SM, Ellena JF, Cafiso DS, Kadokura H, Beckwith J, Bushweller JH (2008) NMR solution structure of the integral membrane enzyme DsbB: functional insights into DsbB-catalyzed disulfide bond formation. Mol Cell 31:896–908
Roosild TP, Greenwald J, Vega M, Castronovo S, Riek R, Choe S (2005) NMR structure of Mistic, a membrane-integrating protein for membrane protein expression. Science 307:1317–1321
Cierpicki T, Liang B, Tamm LK, Bushweller JH (2006) Increasing the accuracy of solution NMR structures of membrane proteins by application of residual dipolar couplings. High-resolution structure of outer membrane protein. A J Am Chem Soc 128:6947–6951
Fernandez C, Wider G (2003) TROSY in NMR studies of the structure and function of large biological macromolecules. Curr Opin Struct Biol 13:570–580
Baker KA, Tzitzilonis C, Kwiatkowski W, Choe S, Riek R (2007) Conformational dynamics of the KcsA potassium channel governs gating properties. Nat Struct Mol Biol 14:1089–1095
Tugarinov V, Muhandiram R, Ayed A, Kay LE (2002) Four-dimensional NMR spectroscopy of a 723-residue protein: chemical shift assignments and secondary structure of malate synthase g. J Am Chem Soc 124:10025–10035
Fernandez C, Hilty C, Wider G, Guntert P, Wuthrich K (2004) NMR structure of the integral membrane protein OmpX. J Mol Biol 336:1211–1221
Oxenoid K, Chou JJ (2005) The structure of phospholamban pentamer reveals a channel-like architecture in membranes. Proc Natl Acad Sci U S A 102:10870–10875
Hilty C, Wider G, Fernandez C, Wuthrich K (2004) Membrane protein–lipid interactions in mixed micelles studied by NMR spectroscopy with the use of paramagnetic reagents. Chembiochem 5:467–473
Beel AJ, Mobley CK, Kim HJ, Tian F, Hadziselimovic A, Jap B, Prestegard JH, Sanders CR (2008) Structural studies of the transmembrane C-terminal domain of the amyloid precursor protein (APP): does APP function as a cholesterol sensor? Biochemistry 47:9428–9446
Mascioni A, Eggimann BL, Veglia G (2004) Determination of helical membrane protein topology using residual dipolar couplings and exhaustive search algorithm: application to phospholamban. Chem Phys Lipids 132:133–144
Cierpicki T, Bushweller JH (2004) Charged gels as orienting media for measurement of residual dipolar couplings in soluble and integral membrane proteins. J Am Chem Soc 126:16259–16266
Kamen DE, Cahill SM, Girvin ME (2007) Multiple alignment of membrane proteins for measuring residual dipolar couplings using lanthanide ions bound to a small metal chelator. J Am Chem Soc 129:1846–1847
Bermel W, Bertini I, Felli IC, Lee YM, Luchinat C, Pierattelli R (2006) Protonless NMR experiments for sequence-specific assignment of back bone nuclei in unfolded proteins. J Am Chem Soc 128:3918–3919
Zhang Q, Atreya HS, Kamen DE, Girvin ME, Szyperski T (2008) GFT projection NMR based resonance assignment of membrane proteins: application to subunit C of E. coli F(1)F(0) ATP synthase in LPPG micelles. J Biomol NMR 40:157–163
Hwang PM, Choy WY, Lo EI, Chen L, Forman-Kay JD, Raetz CR, Prive GG, Bishop RE, Kay LE (2002) Solution structure and dynamics of the outer membrane enzyme PagP by NMR. Proc Natl Acad Sci U S A 99:13560–13565
Hwang PM, Bishop RE, Kay LE (2004) The integral membrane enzyme PagP alternates between two dynamically distinct states. Proc Natl Acad Sci U S A 101:9618–9623
Hwang PM, Kay LE (2005) Solution structure and dynamics of integral membrane proteins by NMR: a case study involving the enzyme PagP. Methods Enzymol 394:335–350
Li C, Gao P, Qin H, Chase R, Gor’kov PL, Brey WW, Cross TA (2007) Uniformly aligned full-length membrane proteins in liquid crystalline bilayers for structural characterization. J Am Chem Soc 129:5304–5305
De Angelis AA, Opella SJ (2007) Bicelle samples for solid-state NMR of membrane proteins. Nat Protoc 2:2332–2338
Park SH, Loudet C, Marassi FM, Dufourc EJ, Opella SJ (2008) Solid-state NMR spectroscopy of a membrane protein in biphenyl phospholipid bicelles with the bilayer normal parallel to the magnetic field. J Magn Reson 193:133–138
Mahalakshmi R, Marassi FM (2008) Orientation of the Escherichia coli outer membrane protein OmpX in phospholipid bilayer membranes determined by solid-state NMR. Biochemistry 47:6531–6538
Marassi FM, Opella SJ (2003) Simultaneous assignment and structure determination of a membrane protein from NMR orientational restraints. Protein Sci 12:403–411
De Angelis AA, Howell SC, Nevzorov AA, Opella SJ (2006) Structure determination of a membrane protein with two trans-membrane helices in aligned phospholipid bicelles by solid-state NMR spectroscopy. J Am Chem Soc 128:12256–12267
Achuthan S, Asbury T, Hu J, Bertram R, Cross TA, Quine JR (2008) Continuity conditions and torsion angles from ssNMR orientational restraints. J Magn Reson 191:24–30
Sinha N, Grant CV, Park SH, Brown JM, Opella SJ (2007) Triple resonance experiments for aligned sample solid-state NMR of (13)C and (15)N labeled proteins. J Magn Reson 186:51–64
Li Y, Berthold DA, Gennis RB, Rienstra CM (2008) Chemical shift assignment of the transmembrane helices of DsbB, a 20-kDa integral membrane enzyme, by 3D magic-angle spinning NMR spectroscopy. Protein Sci 17:199–204
Huang L, McDermott AE (2008) Partial site-specific assignment of a uniformly (13)C, (15)N enriched membrane protein, light-harvesting complex 1 (LH1), by solid state NMR. Biochim Biophys Acta 1777:1098–1108
Lorch M, Fahem S, Kaiser C, Weber I, Mason AJ, Bowie JU, Glaubitz C (2005) How to prepare membrane proteins for solid-state NMR: a case study on the alpha-helical integral membrane protein diacylglycerol kinase from E. coli. Chembiochem 6:1693–1700
Pauli J, van Rossum B, Forster H, de Groot HJ, Oschkinat H (2000) Sample optimization and identification of signal patterns of amino acid side chains in 2D RFDR spectra of the alpha-spectrin SH3 domain. J Magn Reson 143:411–416
Manolikas T, Herrmann T, Meier BH (2008) Protein structure determination from 13C spin-diffusion solid-state NMR spectroscopy. J Am Chem Soc 130:3959–3966
Pauli J, Baldus M, van Rossum B, de Groot H, Oschkinat H (2001) Back bone and side-chain 13C and 15N signal assignments of the alpha-spectrin SH3 domain by magic angle spinning solid-state NMR at 17.6 Tesla. Chembiochem 2:272–281
Castellani F, van Rossum B, Diehl A, Schubert M, Rehbein K, Oschkinat H (2002) Structure of a protein determined by solid-state magic-angle-spinning NMR spectroscopy. Nature 420:98–102
Lange A, Becker S, Seidel K, Giller K, Pongs O, Baldus M (2005) A concept for rapid protein-structure determination by solid-state NMR spectroscopy. Angew Chem Int Ed Engl 44:2089–2092
Malia TJ, Wagner G (2007) NMR structural investigation of the mitochondrial outer membrane protein VDAC and its interaction with antiapoptotic Bcl-xL. Biochemistry 46:514–525
Assadi-Porter FM, Tonelli M, Maillet E, Hallenga K, Benard O, Max M, Markley JL (2008) Direct NMR detection of the binding of functional ligands to the human sweet receptor, a heterodimeric family 3 GPCR. J Am Chem Soc 130:7212–7213
Etzkorn M, Kneuper H, Dunnwald P, Vijayan V, Kramer J, Griesinger C, Becker S, Unden G, Baldus M (2008) Plasticity of the PAS domain and a potential role for signal transduction in the histidine kinase DcuS. Nat Struct Mol Biol 15:1031–1039
Ader C, Schneider R, Hornig S, Velisetty P, Wilson EM, Lange A, Giller K, Ohmert I, Martin-Eauclaire MF, Trauner D, Becker S, Pongs O, Baldus M (2008) A structural link between inactivation and block of a K+ channel. Nat Struct Mol Biol 15:605–612
Lange A, Giller K, Hornig S, Martin-Eauclaire MF, Pongs O, Becker S, Baldus M (2006) Toxin-induced conformational changes in a potassium channel revealed by solid-state NMR. Nature 440:959–962
Etzkorn M, Martell S, Andronesi OC, Seidel K, Engelhard M, Baldus M (2007) Secondary structure, dynamics, and topology of a seven-helix receptor in native membranes, studied by solid-state NMR spectroscopy. Angew Chem Int Ed Engl 46:459–462
Seidel K, Andronesi OC, Krebs J, Griesinger C, Young HS, Becker S, Baldus M (2008) Structural characterization of Ca(2+)-ATPase-bound phospholamban in lipid bilayers by solid-state nuclear magnetic resonance (NMR) spectroscopy. Biochemistry 47:4369–4376
Traaseth NJ, Verardi R, Torgersen KD, Karim CB, Thomas DD, Veglia G (2007) Spectroscopic validation of the pentameric structure of phospholamban. Proc Natl Acad Sci U S A 104:14676–14681
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Montaville, P., Jamin, N. (2010). Determination of Membrane Protein Structures Using Solution and Solid-State NMR. In: Lacapère, JJ. (eds) Membrane Protein Structure Determination. Methods in Molecular Biology, vol 654. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-60761-762-4_14
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