Figure 1.
Overall comparison of native HRV2 and the derived B-particle.
(A) Cartoon diagrams of the native (blue) and the 80S subviral B-particle (orange). Only half of each capsid shell is represented, as a ∼80 Å slab, to illustrate the expansion of the 80S particle with respect to the native virion. The positions of one three-fold and one five-fold axis of the icosahedral particle are indicated with black symbols. Superimposition of the β-barrels of VP1 (B), VP2 (C) and VP3 (D) capsid proteins in the two structures (native capsid in blue, 80S particle in orange). The structural motifs corresponding to the most relevant differences are indicated.
Table 1.
Data collection and refinement statistics.
Figure 2.
Side by side comparison of the VP1 structures from the native virion (A) and the 80S particle (B), showing the hinge movement displacing the αA helix and the C-terminus of VP1 away from the VP1 β-barrel. The secondary structural elements are indicated on the 80S VP1 representation. The angle formed by the position of the Cα atoms from residues Ser1130, Lys1243 and Ala1105 and its value is also shown in both cases. (C) Schematic representation of the protomer expansion in the transition from the native virion (left) to the 80S particle (right). The hinge movement of VP1 (shown in blue) induces a concerted displacement of VP2 and VP3 (in green and red, respectively). To facilitate the comparison, the outer limits of the native protomer are also displayed in the 80S figure as thin lines. (D) Surface representation of the 80S protomer, seen from the interior of the particle, following the five-fold axis. VP1 is shown in blue, VP2 in green and VP3 in red. For clarity, only the Cα are displayed. The position of the symmetry axes is indicated with black symbols (E) Same view as in (D), with each residue colored according to the displacement suffered for its Cα in the transition from the native capsid to the 80S particle. The color scale, indicated as a bar, covers distances from 0 Å (in dark blue) to 5.5 Å or more (in red). The distances were calculated from a superposition of the native and the 80S protomers, using the VP1 β-barrel as a guide. The disulfide bridge linking Cys2229 and Cys3120 is shown as purple sticks and indicated with an arrowhead.
Figure 3.
Changes around the five-fold channels.
Lateral view of the five-fold axis in the 80S particle (A) and the native capsid (B) of HRV2. VP1 protein is shown in blue, VP3 in red and VP4 in yellow. For clarity, only two subunits of VP1 are displayed. In the native structure, the pocket factor is displayed as orange sticks and the N-terminal α-helix (residues 1001 to 1013, not present in the crystal structure of HRV2) has been modeled according to its position in the structure of HRV16 and is shown in grey. The close-ups at the right show a top view of the five-fold axes, with residues His1138 and His1173 from the five symmetry related VP1 subunits displayed as yellow sticks. (C) Close-up of the N-terminal region of VP1 from the HRV2 native structure, seen from the inside of the virion. VP1 is shown in blue, VP2 in green and VP3 in red; the first residue seen in the 80S density map (Arg1062) is indicated with an asterisk and from then on VP1 is colored in cyan. The side chains of the residues involved in acid-labile interactions stabilizing the conformation of the VP1 N-terminus are shown as sticks and explicitly labeled.
Figure 4.
Structural changes in the hydrophobic pocket of VP1.
Stereo views of the VP1 hydrophobic pocket in the empty capsid (A) and in the native HRV2 (B). For clarity, only some of the structural elements forming the pocket are shown. VP1 is represented as a blue cartoon and the relevant residues are shown as blue sticks and explicitly labeled. In (A), the averaged electron density map (contoured at 1.0 σ) is shown as a dark blue mesh around the GH loop. In (B), the pocket factor (a 12-carbon fatty acid) is depicted as orange sticks. The pocket cavity, in the 80S particle, adopts a closed conformation and the side chain of Met1213 has extended, crossing the binding site of the pocket factor. In the native capsid, the polar head group of the pocket factor makes hydrogen bonds with the main chain nitrogen from Leu1101 and Nε of Gln1102 (located in the CD loop) and with the Nγ of Asn1211 (GH loop, just before the H ß-strand). Conversely, in the 80S empty capsid, Asn1211 interacts with the main chain of Tyr1192 (located at the other end of the same GH loop, in the proximity of the hinge pivotal center). This allows Met1213 to occupy the position of the polar head group of the fatty acid, via interaction with residues Ile1099 and Tyr1191.
Figure 5.
Formation of pores in the 80S empty particle.
Surface representation of a pentamer from the 80S particle (A) and the native capsid (B), seen from the outside and along the five-fold symmetry axis. In both figures, VP1 is shown in blue, VP2 in green and VP3 in red. The residues located at the outer opening of the pores in the 80S structure (Met1104, Ala1105, Gln1106, Tyr1159 and Gln1162) and the corresponding residues in the native capsid are painted in yellow. In (A), the location of one pore is explicitly highlighted with a yellow square. (C) Lateral view of the pentamer region highlighted in (A), represented as surface and slabbed to show the pore that completely crosses the capsid from the canyon floor to the capsid interior. The side chains of residues located along the pore walls are shown as sticks in blue (VP1) and red (VP3). The N-terminus of the 80S VP1 structure is also shown as sticks and the residues located at the base of the pore (Arg1062, Asp1063 and Glu1064) are labeled. The side chain of residue Arg1062 has been modeled from the native capsid structure. For orientation purposes, the direction of the five-fold symmetry axis is indicated with a line and a black symbol at the upper-left corner.
Figure 6.
Opening of the two-fold channels in the 80S empty capsid.
Contacts between pentamers along the two-fold and three-fold axes in the 80S empty particle (A) as compared to those present in the native HRV2 capsid (B). The capsid proteins VP1 (blue), VP2 (green) and VP3 (red) are depicted as ribbons. The position of the symmetry axes is indicated in (B). (C) Close-up view of the channel formed at the two-fold axes in the 80S particle, seen from the interior of the capsid. The VP2 and VP3 subunits surrounding the channel are represented as cartoons and the side chains of VP2 exposed at the interface are depicted as sticks. The lines crossing the cavity correspond to distances of 10.2 Å (a) and 30.5 Å (b). (D) Charge distribution on the walls of the channel at the two-fold axes. In order to show the surface of the longitudinal walls, a side view of the channel (perpendicular to the direction of the symmetry axis) has been slabbed. The surface is represented with the electrostatic potential, colored in blue and red for positive and negative charges, respectively. The relevant residues are shown as sticks and labeled. The direction of the two-fold symmetry axis is also indicated with a line and a black symbol.