1. Volume 31, Issue 5, Pages 749-761 (September 2008)
Insights into Virus Evolution and Membrane Biogenesis from the Structure of the Marine Lipid-Containing Bacteriophage PM2 
Nicola G.A. Abrescia, Jonathan M. Grimes, Hanna M. Kivelä, Rene Assenberg, Geoff C. Sutton, Sarah J. Butcher, Jaana K.H. Bamford, Dennis H. Bamford, David I. Stuart 
Molecular Cell 
Volume 31, Issue 5, Pages (September 2008)
DOI: /j.molcel
Copyright © 2008 Elsevier Inc. Terms and Conditions

2. Figure 1 Structural Components of PM2
(A) (Left) Electron density (1σ) of the virus, cut open to reveal the lipid bilayer and enclosed genome (0.25σ, inner surface white). (Center) Cα trace of P1 in the virus map (0.8σ, blue) with SeMet difference Fourier (4σ, red), color coded by domain; red, the modeled penton domain (residues 1–85); dark and light orange, respectively, the stem (residues 86–165 and 325–335) and receptor binding domains (residues 166–324). Green spheres denote methionine positions. (Right) 2Fo − Fc electron density of the receptor-binding domain calcium-binding site (cyan and red spheres show the calcium ion and coordinating waters).
(B) (Left) Major capsid protein P2 trimers within the icosahedral asymmetric unit color coded in yellow, green, cyan, and blue. P1 color coded as above. The white triangle delineates a virus facet, and numbers indicate icosahedral symmetry axes. (Center) Cα trace of P2 (yellow) atomic structure fitted in the virus map (1σ, blue), and SeMet difference Fourier (4σ, red). (Right) 2Fo − Fc electron density for P2 (all 269 residues are visible in the electron density).
(C) (Left) 240 P3 subunits and 60 P6 subunits (dark magenta and gold, respectively) wrap around the membrane vesicle (light cyan). (Center and right) Cα models for P6 and a dimer of P3 fitted in the corresponding virus electron density (0.5σ, blue); red, SeMet peak corresponding to Met52 in P3 (3.2σ).
Molecular Cell  , DOI: ( /j.molcel )
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3. Figure 2
Protein P2
(A) Stereo view of superimposed Cα traces of P2 (jelly roll V1, residues 1–141 in blue; jelly roll V2, residues 142–269 in light blue) and STIV major capsid protein (yellow). Blue and red spheres denote the N and the C termini; also in red are C-terminal residues 264–269. The calcium ion between V1 and V2 is colored cyan.
(B) Structure-based phylogenetic trees of (left) the coat proteins of members of the PRD1-adenovirus lineage using SHP (Stuart et al., 1979) and PHYLIP (Felsenstein, 1989) and (right) individual V1 and V2 domains. V1 and V2 cluster separately (blue and light blue areas) except for PM2.
Molecular Cell  , DOI: ( /j.molcel )
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4. Figure 3
Protein P1
(A) P1 vertex assembly viewed (left) orthogonal to the 5-fold axis (one monomer colored as in Figure 1A) and (right) along the 5-fold axis from above the virus. Calcium ions are in cyan.
(B) (Top) Stereo view of the superimposition of the P1 receptor-binding domain (orange) with Aga16B-CBM6 (violet) (2.7 Å rmsd, 104 Cαs equivalent). Cyan and pink spheres mark the calcium ions in P1 and Aga16b-CBM6. (Center) Schematic of the arrangement of strands forming the interlocked collar comprising five stem domains. The view is along the 5-fold axis, and the β strands, shown as circles, are labeled BIDG and CHEF, conventionally for viral jelly rolls. One of the five subunits is colored orange. (Bottom) Comparison of PM2, PRD1, and adenovirus penton proteins.
(C) Structure-based phylogenetic tree of cell-attachment viral protein domains; Aga16B-CBM6 is a carbohydrate-binding protein from the marine bacterium Saccharophagus degradans (colored as in Figure 3B, top; in parenthesis is the corresponding PDB ID code). The matrix with the evolutionary distances was obtained using SHP (Stuart et al., 1979) and represented using PHYLIP (Felsenstein, 1989).
(D) Phage adsorption inhibition in the presence of the purified P1 and the receptor-binding domain of P1 (P1-RBD, residues 159–335). Phage adsorption to ER72M2 cells was measured after 10 min of adsorption in the presence of increasing amounts of protein. As a nonspecific control, the recombinant PM2 capsid protein P2 had no inhibitory effects on PM2 entry. The inset shows the purified protein P1 and the receptor-binding fragment used in the adsorption inhibition tests (Coomassie blue-stained polyacrylamide gel). The molecular mass markers are indicated on the left.
Molecular Cell  , DOI: ( /j.molcel )
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5. Figure 4 PM2 Architecture
(Top center) Schematic of the PM2 capsid (gray) and membrane (cyan). Hexagons and triangles represent pseudohexameric P2 trimers and red pentagons the P1 penton domains. (Top right) Equivalent representation of bacteriophage PRD1. The black lines indicate the pseudo T numbers (PM2, h = 4, k = 1; PRD1, h = 5, k = 0, where T = h2 + hk + k2; PM2 and PRD1 are not drawn to the same scale). The gray oval defines the portion enlarged in the central diagram where trimers are represented as color-coded hexagons, 2- and 3-fold axes as red ovals and triangles respectively, and pseudo 3-folds as blue triangles. The dark magenta helix-turn-helix represents protein P3 with the C-terminal transmembrane helices as a circle. The gold circle and helix denote P6. For clarity, P3 and P6 are depicted above the capsid. The close-up linked by straight zoom lines is viewed as in the main diagram, and square close-ups (A–C) indicate tangential views showing the P3/P6 complex with the transmembrane helices inserting along the line joining the 5-folds (the electron density for the headgroups [0.4σ, cyan] and acyl-chain region [−1.3σ, light red] shadowed in gray). (Right) Close-ups describe the organization of the P2 and P3 trimers at the icosahedral 3-fold axis; cyan spheres depict the calcium ions.
Molecular Cell  , DOI: ( /j.molcel )
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6. Figure 5
Proteins P3 and P6
(A and B) Stereo pictures of contiguous views of a portion of Cα traces of a P3 dimer (magenta) and P6 (gold) fitted in the virus map (0.7σ, blue), with SeMet difference Fourier (3.2σ, red) viewed orthogonally to the virus surface.
Molecular Cell  , DOI: ( /j.molcel )
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7. Figure 6 Membrane and Genome Architecture
(A) (Left) Slice through the PM2 electron density. The capsid is blue (0.8σ), lipid headgroups cyan (0.25σ), acyl-chain region light red (−1.3σ), and supercoiled DNA green (0.25σ). (Right) Icosahedral electron density profile. Distances measured from the particle center along the icosahedral 3-fold axis. IL and OL mark the inner and outer membrane leaflets, respectively.
(B) Cartoon of PM2 membrane vesicle assembly. (1) Dimers of protein P3 (magenta) and a monomer of protein P6 (gold) anchored via transmembrane helices (data not shown) on a patch of bacterial membrane. (2) Independent P3 dimers interact with monomeric P6 forming the scaffold building block. (3) Three building blocks come together by interaction of the P3 α1 helices to form a subassembly corresponding to an icosahedral facet. (4) P6 molecules of two independent subassemblies interact, facilitated by interaction with the supercoiled DNA genome via P6 transmembrane helices (and possibly further components such as P4). (5) This interaction generates a torque across the membrane via the P6 helices (depicted as small gold-colored rectangles), driving the curvature of the membrane. (6) Recruitment of further P6-P3 subassemblies to the condensed DNA genome leads to a correctly sized lipid vesicle coated with P3 and P6, on which the outer protein capsid assembles.
Molecular Cell  , DOI: ( /j.molcel )
Copyright © 2008 Elsevier Inc. Terms and Conditions