Backbone structure of the infectious ε15 virus capsid revealed by electron cryomicroscopy

A half- century after the determination of the first three-dimensional crystal structure of a protein(1), more than 40,000 structures ranging from single polypeptides to large assemblies have been reported(2). The challenge for crystallographers, however, remains the growing of a diffracting crystal. Here we report the 4.5- angstrom resolution structure of a 22-MDa macromolecular assembly, the capsid of the infectious epsilon15 ( epsilon 15) particle, by single-particle electron cryomicroscopy. From this density map we constructed a complete backbone trace of its major capsid protein, gene product 7 ( gp7). The structure reveals a similar protein architecture to that of other tailed double- stranded DNA viruses, even in the absence of detectable sequence similarity(3,4). However, the connectivity of the secondary structure elements ( topology) in gp7 is unique. Protruding densities are observed around the two- fold axes that cannot be accounted for by gp7. A subsequent proteomic analysis of the whole virus identifies these densities as gp10, a 12- kDa protein. Its structure, location and high binding affinity to the capsid indicate that the gp10 dimer functions as a molecular staple between neighbouring capsomeres to ensure the particle's stability. Beyond epsilon 15, this method potentially offers a new approach for modelling the backbone conformations of the protein subunits in other macromolecular assemblies at near- native solution states.