High resolution 3D structures by electron cryomicroscopy
First, many membrane proteins, including most G-protein-coupled receptors, are relatively unstable after purification in detergent: for this class of membrane proteins, Chris Tate and his colleagues in the LMB have recently developed the method of conformational stabilisation (Serrano-Vega et al (2008) PNAS, 105, 877-882) by combining a number of thermostabilising mutants to create much more stable molecules. This has made it possible to tackle structures that were practically impossible previously.
A second problem is that X-ray or electron crystallography needs well-ordered crystals that diffract to high resolution. Since the recent introduction of direct electron detectors and improved computer programs, the resolution of single particle electron cryomicroscopy (cryoEM) has been greatly improved to the extent that it is now a serious alternative to crystallography. In many but not all cases, atomic structures can now be obtained using single particle cryoEM. We are therefore working to improve the imaging methods, the computer programs for image processing, and the efficiency of electron detectors, with the goal of realising the full potential of cryoEM for analysis of single particle structure at atomic resolution.
- Vinothkumar, K.R. and Henderson, R. (2010)
Structures of Membrane Proteins.
Quarterly Reviews of Biophysics 43: 65-158.
- Henderson, R., Chen, S., Chen, J.Z., Grigorieff, N., Passmore, L.A., Ciccarelli, L., Rubinstein, J.L., Crowther, R.A., Stewart, P.L. and Rosenthal, P.B. (2011)
Tilt-pair analysis of images from a range of different specimens in single particle electron cryomicroscopy.
J Mol Biol 413: 1028-1046.
- Chen, S., McMullan, G., Faruqi, A.R., Murshudov, G.N., Short, J.M., Scheres, S.H. & Henderson, R. (2013)
High-resolution noise substitution to measure overfitting and validate resolution in 3D structure determination by single particle electron cryomicroscopy.
Ultramicroscopy 135: 24-35.
- Wasi Faruqi
- Greg McMullan
- Vinothkumar Kutti Ragunath
- Daria Slowik