In polyomaviruses the pentameric capsomers are interlinked by the long C-terminal arm of the structural protein VP1. The T‫7؍‬ icosahedral structure of these viruses is possible due to an intriguing adaptability of this linker arm to the different local environments in the capsid. To explore the assembly process, we have compared the structure of two virus-like particles (VLPs) formed, as we found, in a calcium-dependent manner by the VP1 protein of human polyomavirus BK. The structures were determined using electron cryomicroscopy (cryo-EM), and the three-dimensional reconstructions were interpreted by atomic modeling. In the small VP1 particle, 26.4 nm in diameter, the pentameric capsomers form an icosahedral T‫1؍‬ surface lattice with meeting densities at the threefold axes that interlinked three capsomers. In the larger particle, 50.6 nm in diameter, the capsomers form a T‫7؍‬ icosahedral shell with three unique contacts. A folding model of the BKV VP1 protein was obtained by alignment with the VP1 protein of simian virus 40 (SV40). The model fitted well into the cryo-EM density of the T‫7؍‬ particle. However, residues 297 to 362 of the C-terminal arm had to be remodeled to accommodate the higher curvature of the T‫1؍‬ particle. The loops, before and after the C-terminal short helix, were shown to provide the hinges that allowed curvature variation in the particle shell. The meeting densities seen at the threefold axes in the T‫1؍‬ particle were consistent with the triple-helix interlinking contact at the local threefold axes in the T‫7؍‬ structure.The BK virus (BKV) is a human virus belonging to the Polyomaviridae family. It is a nonenveloped virus (ϳ50.0 nm in diameter) with a circular double-stranded DNA genome (ϳ5 kb). The capsid has icosahedral symmetry and is built of 72 capsomers that are all pentamers of the protein VP1 arranged in a Tϭ7 icosahedral lattice (21). All known polyomaviruses have three structural proteins (VP1, VP2, and VP3), of which VP1 is the major capsid protein. Overall amino acid sequence homology between BKV and the other human polyomavirus, JCV, is 75%, and that with the simian polyomavirus (SV40) is 69% (9). In the VP1 protein the sequence similarity rises to 77% and 74% for the JCV and SV40, respectively (35). Due to the high similarity, the solved atomic structure of VP1 of SV40 provides us a template to create a model of the BKV VP1 protein folding.The structures of the SV40 and murine polyomavirus have been determined and show similar features to that seen in the BKV (1, 12). The VP1 pentamer of SV40 and murine polyomavirus is built as a ring of five ␤-barrel-shaped VP1 monomers, tightly linked by interacting loops between the framework of ␤-strands (22,33,34,40). The C-terminal subdomain of each VP1 monomer "invades" a neighboring pentamer, thereby tying the pentamers together in the virion shell. There are six unique monomers building up the capsid (monomer ␣, ␣Ј, and ␣Љ at the local threefold; ␤, ␤Ј around the icosahedral threefold, and ␥ at the twofold) (34). The major s...

Semliki Forest virus is among the prototypes for Class II virus fusion and targets the endosomal membrane. Fusion protein E1 and its envelope companion E2 are both anchored in the viral membrane and form an external shell with protruding spikes. In acid environments, mimicking the early endosomal milieu, surface epitopes in the virus rearrange along with exposure of the fusion loop. To visualize this transformation into a fusogenic stage, we determined the structure of the virus at gradually lower pH values. The results show that while the fusion loop is available for external interaction and the shell and stalk domains of the spike begin to deteriorate, the E1 and E2 remain in close contact in the spike head. This unexpected observation points to E1 and E2 cooperation beyond the fusion loop exposure stage and implies a more prominent role for E2 in guiding membrane close encounter than has been earlier anticipated.Membrane fusion is important in biology to allow exchange of materials between compartments enclosed by lipid bilayer membranes. Naturally, viruses utilize such ways of genome delivery. Two different mechanisms of virus fusion are observed: the Class I mechanism, seen in viruses where the fusion protein structure is dominated by ␣-helix folding, like the influenza and the HIV, and the Class II mechanism, represented by viruses where the envelope proteins are dominated by ␤-sheet structures like the Tick Borne Encephalitis virus and the Semliki Forest virus (SFV) 2 in the Flavi and Toga virus families, respectively (1, 2).SFV infects its host cells by receptor-mediated endocytosis and fusion within acidic endosomes. It has been demonstrated that the membrane fusion of SFV is strictly dependent on the exposure of the virus to a low pH (3-5) and on the presence of cholesterol in the target membrane (6). Experimental mimicking of the endosomal acidification has shown that low pH triggers a series of conformational changes in the virus spikes (7-10). Mild acidification leads to exposure of the fusion peptide at the virion surface (11). The moderately hydrophobic fusion peptide loop can then interact with the target membrane and initiate fusion (12, 13). Optimal fusion kinetics of SFV has been observed at pH 5.5, while the pH threshold for fusion activation is reported to be around pH 6.2 (3, 5).Early data on the alphavirus imply that the fusion process involves a low pH-induced conformational change in the envelope whereby homotrimers of E1 are formed (9,10,14). This is supported by recent studies on the soluble ectodomain of the fusion protein, E1*. The crystal structure of E1* reveals an elongated molecule, mainly folded into ␤-sheets and interconnecting loops (15, 16). The structure resembles that of the TBE glycoprotein E ectodomain (17). The molecule comprises three domains; a ␤-sheet-folded domain I (DI) located in the central part of the molecule inbetween the projecting domain II (DII), carrying the putative fusion peptide loop (18,19) at the distal end, and the immunoglobulin-like domain III (DIII...

The structures of the double-shelled rice dwarf virus and of its single-shell core have been determined by cryoelectron microscopy and image reconstruction. The core carries a prominent density located at each of the icosahedral faces of its T = 1 lattice. These protrusions are formed by outer shell trimers, tightly inserted at the threefold positions of the core. Such configuration of the core may guide the assembly of the outer shell, aided by lateral interactions between its subunits, into a T = 13 lattice. The organization of the phytoreovirus capsid elucidates for the first time a general model for assembling two unique T numbers of quasi-equivalence.

The capsid structures of particles of Rice dwarf virus that consisted of different components, namely, intact particles, empty particles lacking the 12 segments of double-stranded RNA (dsRNA), and virus-like particles composed of only the P3 core and P8 outer capsid proteins, generated with a baculovirus gene-expression system, were determined by cryo-electron microscopy. Combining the results with those of biochemical analysis, we assigned proteins of the transcriptional machinery and dsRNA to density clusters around the 5-fold axes and along the radial concentric layers, respectively. P7 protein, a component of the transcriptional machinery, was assigned to the outermost region of the density clusters. The density connecting the transcription complex to the outermost RNA densities implied interactions between the dsRNA and the P7 protein. Our structural analysis and the non-specific nucleic acid-binding activity of P7 explain the spiral organization of dsRNA around the 5-fold axis.

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