Determination of Molecular Structures of HIV Envelope Glycoproteins using Cryo-Electron Tomography and Automated Sub-tomogram Averaging

JoVE 2770 12/01/2011

Joel R. Meyerson^1,2, Tommi A. White¹, Donald Bliss³, Amy Moran³, Alberto Bartesaghi¹, Mario J. Borgnia¹, M. Jason V. de la Cruz¹, David Schauder¹, Lisa M. Hartnell¹, Rachna Nandwani^1,4, Moez Dawood⁵, Brianna Kim⁶, Jun Hong Kim⁷, John Sununu⁸, Lisa Yang⁹, Siddhant Bhatia¹⁰, Carolyn Subramaniam¹, Darrell E. Hurt¹¹, Laurent Gaudreault¹², Sriram Subramaniam¹

¹Laboratory of Cell Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, ²The Medical Research Council Mitochondrial Biology Unit, University of Cambridge, ³National Library of Medicine, National Institutes of Health, ⁴Massachusetts Institute of Technology, ⁵William Fremd High School, ⁶University of Virginia, ⁷Duke University, ⁸Yale University, ⁹University of Notre Dame, ¹⁰Washington University in St. Louis, ¹¹Bioinformatics and Computational Biosciences Branch, National Institute of Allergy and Infectious Diseases, National Institutes of Health, ¹²Thomas Jefferson High School for Science and Technology

The protocol describes a high-throughput approach to determining structures of membrane proteins using cryo-electron tomography and 3D image processing. It covers the details of specimen preparation, data collection, data processing and interpretation, and concludes with the production of a representative target for the approach, the HIV-1 Envelope glycoprotein. These computational procedures are designed in a way that enables researchers and students to work remotely and contribute to data processing and structural analysis.