title: Visualization of the target-membrane-inserted fusion protein of Semliki Forest virus by combined electron microscopy and crystallography.
abstract: Semliki Forest virus enters cells by receptor-mediated endocytosis. The acidic environment of the endosome triggers a membrane fusion reaction that is mediated by the E1 glycoprotein. During fusion, E1 rearranges from an E1/E2 heterodimer to a highly stable, membrane-inserted E1 homotrimer (E1HT). In this study, we analyzed E1HT by a combination of electron cryomicroscopy, electron crystallography of negatively stained 2D crystals, and fitting of the available X-ray structure of the monomeric E1 ectodomain into the resulting 3D reconstruction. The visualized E1HT reveals that the ectodomain has reoriented vertically and inserted the distal tip of domain II into the lipid bilayer. Our data allow the visualization of a viral fusion protein inserted in its target membrane and demonstrate that insertion is a cooperative process, resulting in rings composed of five to six homotrimers.
mesh_terms: D000818:Animals; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism* / Q000821:virology; D002499:Centrifugation, Density Gradient; D006224:Cricetinae; D018360:Crystallography, X-Ray; D019281:Dimerization; D004705:Endocytosis; D005821:Genetic Techniques; D007091:Image Processing, Computer-Assisted; D008081:Liposomes / Q000378:metabolism; D008561:Membrane Fusion; D008854:Microscopy, Electron; D008958:Models, Molecular; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000378:metabolism*; D012672:Semliki forest virus / Q000235:genetics* / Q000378:metabolism*; D013395:Sucrose / Q000494:pharmacology
chemical_list: D008081:Liposomes; D011993:Recombinant Fusion Proteins; D013395:Sucrose