Membrane Proteins; 2D crystals; Projective Constraint Optimization; Electron Crystallography; Image Processing; Membrane Proteins
Abeyrathne P. D., Arheit M., Kebbel F., Castano-Diez D., Goldie K. N., Chami M., Renault L., Kühlbrandt W., Stahlberg H. (2012), Electron Microscopy Analysis of 2D Crystals of Membrane Proteins, in Comprehensive Biophysics
, 1.19, 277-310.
Abeyrathne P. D., Chami M., Pantelic R. S., Goldie K. N., Stahlberg H. (2012), Preparation of 2D crystals of membrane proteins for high-resolution electron crystallography data collection, in Methods Enzymol
, 481, 25-43.
Gipson B. R., Masiel D. J., Browning N. D., Spence J., Mitsuoka K., Stahlberg H. (2011), Automatic recovery of missing amplitudes and phases in tilt-limited electron crystallography of two-dimensional crystals, in Phys Rev E Stat Nonlin Soft Matter Phys
, 84(1-1), 011916-1-011916-9.
Schenk A. D., Castano-Diez D., Gipson B., Arheit M., Zeng X., Stahlberg H. (2010), 3D reconstruction from 2D crystal image and diffraction data, in Methods Enzymol
, 482(4), 101-129.
Arheit M., Castano-Diez D., Thierry R., Gipson B., Zeng X., Stahlberg H., Automation of Image Processign in Electron Crystallography, in Methods in Molecular Biology
, in press.
Arheit M., Castano-Diez D., Thierry R., Gipson B. R., Zeng X., Stahlberg H., Image Processing for 2D Crystal Images, in Methods in Molecular Biology
, in press.
Arheit M., Castano-Diez D., Thierry R., Abeyrathne P. D., Gipson B. R., Stahlberg H., Merging of Image Data in Electron Crystallography, in Methods in Molecular Biology
, in press.
Membrane proteins are central to health and disease. Electron crystallography can deter-mine the high-resolution structure of membrane proteins from membrane-reconstituted and two-dimensionally crystallized protein. The Transmission Electron Microscope (TEM) is used to study membrane proteins in the lipid membrane environment, which is a powerful tool to study the structure and dynamic of membrane proteins, and of membrane protein complexes that still resist high-resolution 3D crystallization or large-quantity expression efforts.Two-dimensional membrane crystals are imaged by cryo-electron microscopy (cryo-EM). The high-resolution 3D membrane protein structure can then be reconstructed by computer im-age processing. Until recently, the only available software for that task was the MRC software (Crowther et al., 1996). Starting in 2003, my group has created a software system, called 2dx, which embeds the MRC software into a user-friendly graphical user interface, and in addition offers a multitude of additional functions that enable full automation of the processing task, user guidance, interactive documentation and help, and allows automated iterative refinement of the processing results. 2dx received good acceptance in and excellent feedback from the electron crystallography community, and we now count more than 240 external users (as of 1/15/2009).We here propose to extend the 2dx software by new algorithms, to increase resolution and speed of the membrane protein structure determination by electron crystallography, and to en-able high-resolution structure determination also from badly ordered 2D crystals. We will imple-ment a single-particle approach for 3D reconstructions from tilted 2D crystal images, and extend this by a maximum likelihood algorithm. We will develop a hybrid input output algorithm imple-mentation to improve the Z resolution of electron crystallography, and to reduce the need to record images of highly tilted 2D crystals. We will also modernize 2dx, by enabling cluster CPU and cluster GPU computing, and interface it with other systems like Eman2 and IPLT.