Resolving structural details of membrane peptides and proteins at high resolution
Co-workers: Roslin Adamson, Juan Bolivar-Gonzalez, Patricia Dijkman, Peter Fisher, Peter Judge, Marc-Philipp Pfeil, Garrick Taylor, Louic Vermeer, Daniel Yin.
Resolving structural details of membrane receptors is still a major challenge, not least because of the difficulties of expression, crystallization and their size. Since membrane protein secondary structure can be modelled for many cases, we have been devising solid state NMR methods for determining high resolution (sub-Å) details of information-rich sites within membrane receptors (Watts, A., 2005. Nature Reviews Drug Discovery, 4, 555-568). In particular, we have resolved new information about ligand (drug, neurotransmitter or solute) binding sites, and related all this information to functional descriptions.
Our recent focus has been on the neurotensin receptor (NTS1) which we have expressed in E. coli in structural biology amounts (Attrill et al, 2009, J. Express & Purif, 64, 32-38) in a functionally competent form for structural studies, some of which involve single molecule approaches for bionanotechnological and drug design applications.
NTS1 is now available highly purified monodispersed in detergent and in a ligand-binding form. One approach to monitoring ligand binding has been to develop a novel surface plasmon resonance method for tagging the natural ligand, neurotensin (13-mer peptide), to the chip and monitoring protein binding (Harding et al., 2006, Euro. Biophys. J., 35, 709-712). Fluorescently tagged NTS1 has also been used in fluorescence resonance energy transfer methods to resolve long range information of protein-protein signalling (Harding, et al., 2009, Biophsy. J., 96, 964-973).
Publications - 2014
- Liebel M, Schnedermann C, Bassolino G, Taylor G, Watts A, Kukura P (2014) Direct observation of the coherent nuclear response after the absorption of a photon. Physical Review Letters. ; 112, 238301
- Judge, P.J., Taylor, G. F.,Vermeer, L. S. and Watts A. (2014), "Structural Insights from Solid-State NMR into the Function of the Bacteriorhodopsin Photoreceptor Protein" CHAPTER 23 in Advances in Biological Solid-State NMR: Proteins and Membrane-Active Peptides, (eds F. Separovic, A. Naito)
Publications - 2013
- Goddard, A., Dijkman, P., Adamson, R., Watts, A. (2013) Lipid-Dependent GPCR Dimerization. In P. Michael Conn, editors: Receptor-Receptor Interactions, Vol 117, MCB, UK: Academic Press, pp. 341-357.
- Cross, T.A., Murray, D.T. and Watts, A. (2013) Helical Membrane Protein Conformations and their Environment, European Biophysics Journal, Vol. 42, Issue 10, pp 731-755
- Ryan, L., Lamarre, B., Diu, T., Ravi, J., Judge, P., Temple, A., Carr, M., Cerasoli, E., Su, B., Jenkinson, H., Martyna, G., Crain, J., Watts, A., Ryadnov, M. (2013) Anti-antimicrobial peptides: folding-mediated host defense antagonists. Journal of Biological Chemistry, 288, 20162-72.
- Long, A., O'Brien, C., Malhotra, K., Schwall, C., Albert, A., Watts, A., Alder, N. (2013) A Detergent-Free Strategy for the Reconstitution of Active Enzyme Complexes from Native Biological Membranes into Nanoscale Discs. BMC Biotechnology, 13, 41.
- Rakowska, P., Jiang, H., Ray, S., Pyne, A., Lamarre, B., Carr, M., Judge, P., Ravi, J., Gerling, U., Koksch, B., Martyna, G., Hoogenboom, B., Watts, A., Crain, J., Grovenor, C., Ryadnov, M. (2013) Nanoscale imaging reveals laterally expanding antimicrobial pores in lipid bilayers. Proc. Natl. Academ. Sci.(USA), 110, 8918-8923.
- Ding, X., Zhao, X., Watts, A. G-protein-coupled receptor structure, ligand binding and activation as studied by solid-state NMR spectroscopy. Biochem. J. 450, 443-457.
- Seidel, S., Dijkman, P., Lea, W., Bogaart, G., Jerabek-Willemsen, M., Lazic, A., Joseph, J., Srinivasan, P., Baaske, P., Simeonov, A., Katritch, I., Ladbury, J., Schreiber, G., Watts, A., Braun, D., and S. Duhr. (2013) Microscale thermophoresis quantifies biomolecular interactions under previously challenging conditions. Methods 59, 301-315.
Publications - 2012
- Orwick, M., Lovett, J., Graziadei, A., Lindholm, L., Hicks, M., Watts, A.(2012) Detergent-Free Incorporation of a Seven-Transmembrane Receptor Protein into Nanosized Bilayer Lipodisq Particles for Functional and Biophysical Studies. Nano Letters, 12, 4687-4692.
- Watts, A. (2012) Exploiting magnetic resonance spectral anisotropy averaging to gain biological details in biomembranes. Encyclopaedia of Magnetic Resonance – Historical Perspectives (E. D. Becker, Editor) Wiley Interscience.
- Higman, V. and Watts, A. (2012) "Recent Developments in Biomolecular Solid-State NMR", CHAPTER 13 in Recent Developments in Biomolecular NMR, (eds M. Clore and J. Potts).
- Oates, J., Faust, B., Attrill, H., Harding, P., Orwick, M., Watts, A. (2012) The role of cholesterol on the activity and stability of neurotensin receptor 1. BBA - Biomembranes, 1818, 2228-33.
- Goddard, A. and Watts, A. (2012) Contributions of fluorescence techniques to understanding G protein-coupled receptor dimerisation. Biophysical Review, 4, 291-298.
- Goddard, A. and Watts, A. (2012) Regulation of G protein-coupled receptors by palmitoylation and cholesterol. BMC Biology, 10, 27-30.
- Orwick, M., Judge, P., Procek, J., Lindholm, L., Graziadei, A., Engel, A., Grobner, G., Watts, A. (2012) Detergent-free formation and physico-chemical characterization of nanosized lipid-polymer complexes - Lipodisq. Angewante Chemie, 51, 1-6.
- Patil, A., Premaruban, T., Berthoumieu, O., Watts, A., Davis, J. (2012) Engineered Bacteriorhodopsin: A Molecular Scale Potential Switch. Chem. Eur. J., 18, 5632-36.
- Pike, K.J., Kemp, T.F., Takahashi, H., Day, R., Howes, A.P., Kryukov, E.V., MacDonald, J.F., Collis, A.E.C., Bolton, D.R., Wylde, R.J., Orwick, M., Kosuga, K., Clark, A.J., Idehara, T., Watts, A., Smith, G.M., Newton, M.E., Dupree, R., Smith, M.E. (2012) A spectrometer designed for 6.7 and 14.1 T DNP-enhanced solid-state MAS NMR using quasi-optical microwave transmission, J. Mag. Res., 215, 1-9.
- Berthoumieu, O., Patil, A., Wang, X., Aslimovska, L., Davis, J., and Watts, A. (2012) Molecular scale conductance photoswitching in engineered bacteriorhodopsin. Nano Letters, 12, 899–903.
- Patil, A., Premaruban, T., Berthoumieu, O., Watts, A., Davis, J. (2012) Enhanced photocurrent in engineered bacteriorhodopsin monolayer films.J. Phys. Chem. B, 116 , 683–689.
A Fascination with Vision - a video lecture for non-specialists
Figure 1: Substituted imidazo-pyridines are inhibitors of the gastric H+/K+-ATPase. By specific labelling of members of this drug family with NMR visible isotopes, we have been able to define the full conformation of the bound ligand, and suggest mechanism for inhibition from homology modelling with a related protein. (Kim, Watts & Watts, 2005, J. Med. Chem. 48, 7145-7152 and Watts, Watts & Middleton, 2001, J. Biol. Chem., 276, 43197-43204).
Figure 2: The cation-π interaction of acetyl choline, a major brain neurotransmitter, and the ligand gated, nicotinic acetyl choline receptor has been resolved using solid state NMR, giving an insight into the binding mechanism and the residues surrounding the site. (Watts, 2005, Nature Reviews Drug Discovery, 4, 555-568; Williamson et al., 2007, PNAS, 104, 18031-18036)
Figure 3: The way in which retinal is restrained within its binding site in membrane-embedded mammalian rhodopsin, and the structural details of the site, have been resolved for the early activation states of this light-activated GPCR, using high resolution solid state NMR to measure atomic distances within the retinal to high accuracy (+/- 0.2Å). (Spooner, et al., 2004, J. Mol. Biol., 343, 719-730)