Retrovirus envelope protein complex structure in situ studied by cryo-electron tomography

• DOI: 10.1073/pnas.0409178102
• Publication Date: 2005-03-18T04:53:22Z
• Authors: Friedrich Förster; Ohad Medalia; Nathan Zauberman; Wolfgang Baumeister; Deborah Fass
• Keywords: 0301 basic medicine; 03 medical and health sciences; Viral Envelope Proteins; Protein Conformation; Cryoelectron Microscopy; Image Processing, Computer-Assisted; Moloney murine leukemia virus; Tomography; Algorithms; 3. Good health

We used cryo-electron tomography in conjunction with single-particle averaging techniques to study the structures of frozen-hydrated envelope glycoprotein (Env) complexes on intact Moloney murine leukemia retrovirus particles. Cryo-electron tomography allows 3D imaging of viruses in toto at a resolution sufficient to locate individual macromolecules, and local averaging of abundant complexes substantially improves the resolution. The averaging of repetitive features in electron tomograms is hampered by a low signal-to-noise ratio and anisotropic resolution, which results from the “missing-wedge” effect. We developed an iterative 3D averaging algorithm that compensates for this effect and used it to determine the trimeric structure of Env to a resolution of 2.7 nm, at which individual domains can be resolved. Strikingly, the 3D reconstruction is shaped like a tripod in which the trimer penetrates the membrane at three distinct locations ≈4.5 nm apart from one another. The Env reconstruction allows tentative docking of the x-ray crystal structure of the receptor-binding domain. This study thus provides 3D structural information regarding the prefusion conformation of an intact unstained retrovirus surface protein.