Three-Dimensional Structure of the Enveloped Bacteriophage Φ12: An Incomplete T = 13 Lattice Is Superposed on an Enclosed T = 1 Shell

BACKGROUND:Bacteriophage phi12 is a member of the Cystoviridae, a unique group of lipid containing membrane enveloped bacteriophages that infect the bacterial plant pathogen Pseudomonas syringae pv. phaseolicola. The genomes of the virus species contain three double-stranded (dsRNA) segments, and the virus capsid itself is organized in multiple protein shells. The segmented dsRNA genome, the multi-layered arrangement of the capsid and the overall viral replication scheme make the Cystoviridae similar to the Reoviridae. METHODOLOGY/PRINCIPAL FINDINGS:We present structural studies of cystovirus phi12 obtained using cryo-electron microscopy and image processing techniques. We have collected images of isolated phi12 virions and generated reconstructions of both the entire particles and the polymerase complex (PC). We find that in the nucleocapsid (NC), the phi12 P8 protein is organized on an incomplete T = 13 icosahedral lattice where the symmetry axes of the T = 13 layer and the enclosed T = 1 layer of the PC superpose. This is the same general protein-component organization found in phi6 NC's but the detailed structure of the entire phi12 P8 layer is distinct from that found in the best classified cystovirus species phi6. In the reconstruction of the NC, the P8 layer includes protein density surrounding the hexamers of P4 that sit at the 5-fold vertices of the icosahedral lattice. We believe these novel features correspond to dimers of protein P7. CONCLUSIONS/SIGNIFICANCE:In conclusion, we have determined that the phi12 NC surface is composed of an incomplete T = 13 P8 layer forming a net-like configuration. The significance of this finding in regard to cystovirus assembly is that vacancies in the lattice could have the potential to accommodate additional viral proteins that are required for RNA packaging and synthesis

Characterization of recombinant amino-terminal NC4 domain of human collagen IX - Interaction with glycosaminoglycans and cartilage oligomeric matrix protein

The N-terminal NC4 domain of collagen IX is a globular structure projecting away from the surface of the cartilage collagen fibril. Several interactions have been suggested for this domain, reflecting its location and its characteristic high isoelectric point. In an attempt to characterize the NC4 domain in more detail, we set up a prokaryotic expression system to produce the domain. The purified 27.5-kDa product was analyzed for its glycosaminoglycan-binding potential by surface plasmon resonance and solid-state assays. The results show that the NC4 domain of collagen IX specifically binds heparin with a K-d of 0.6 muM, and the full-length recombinant collagen IX has an even stronger interaction with heparin, with an apparent K-d of 3.6 nM. The heparin-binding site of the NC4 domain was located in the extreme N terminus, containing a heparin-binding consensus sequence, whereas electron microscopy suggested the presence of at least three additional heparin-binding sites on full-length collagen IX. The NC4 domain was also shown to bind cartilage oligomeric matrix protein. This interaction and the association of cartilage oligomeric matrix protein with other regions of collagen IX were found to be heparin-competitive. Circular dichroism analyses of the NC4 domain indicated the presence of stabilizing disulfide bonds and a thermal denaturation point of about 80degreesC. The pattern of disulfide bond formation within the NC4 domain was identified by tryptic peptide mass mapping of the NC4 in native and reduced states. A similar pattern was demonstrated for the NC4 domain of full-length recombinant collagen IX

Structure and host-cell interaction of SH1, a membrane-containing, halophilic euryarchaeal virus

The Archaea, and the viruses that infect them, are the least well understood of all of the three domains of life. They often grow in extreme conditions such as hypersaline lakes and sulfuric hot springs. Only rare glimpses have been gained into the structures of archaeal viruses. Here, we report the subnanometer resolution structure of a recently isolated, hypersalinic, membrane-containing, euryarchaeal virus, SH1, in which different viral proteins can be localized. The results indicate that SH1 has a complex capsid formed from single β-barrels, an important missing link in hypotheses on viral capsid protein evolution. Unusual, symmetry-mismatched spikes seem to play a role in host adsorption. They are connected to highly organized membrane proteins providing a platform for capsid assembly and potential machinery for host infection

Structural and Functional Studies of Archaeal Viruses*

Viruses populate virtually every ecosystem on the planet, including the extreme acidic, thermal, and saline environments where archaeal organisms can dominate. For example, recent studies have identified crenarchaeal viruses in the hot springs of Yellowstone National Park and other high temperature environments worldwide. These viruses are often morphologically and genetically unique, with genomes that show little similarity to genes of known function, complicating efforts to understand their viral life cycles. Here, we review progress in understanding these fascinating viruses at the molecular level and the evolutionary insights coming from these studies