The three-dimensional structure of frozen-hydrated bacteriophage phi X174

Abstract

The three-dimensional structure of bacteriophage phi X174 (phi X174) was determined to approximately 2.6 nm resolution from images of frozen-hydrated 114 S particles. The outer surface of phi X174 is characterized by several prominent features: (i) 12 mushroom-shaped caps (approximately 7.1 nm wide x 3.8 nm high) are situated at each of the vertices of the icosahedral virion and extend to a maximum radius of 16.8 nm; (ii) a "collar" of density surrounds the base of each apical cap; and (iii) 20 conical protrusions (approximately 2.3 nm high) lie along the three-fold symmetry axes. The caps have a pentagonal morphology composed of five globular "subunits" and appear to be loosely connected to the underlying capsid. The distribution of the four gene products present in virions (60 copies each of gpF, gpG, and gpJ, and 12 copies of gpH), and the single-stranded DNA (ssDNA) genome cannot be directly discerned in the reconstructed density map, although plausible assignments can be made on the basis of solvent-excluded volume estimates and previous biochemical data. Thus, gpG accounts for most of the mass in the caps; gpH, a presumed cap protein, cannot be identified in part due to the symmetry-averaging procedures, but may be partially located within the interior of the capsid; and gpF and gpJ make up the remainder of the capsid. The genome appears to be less densely packaged inside the capsid compared to many dsDNA viruses whose nucleic acid is arranged in a liquid-crystalline state.

Fig. 1

Frozen-hydrated ϕX174 and polyoma virus (triangle). Apical caps on some particles appear to float as if disconnected from the capsid surface (long arrows) presumably because of defocus-induced Fresnel fringes around the periphery of the particles. Some particles are viewed close to an icosahedral two-fold axis (filled arrow) or have lost most of the ssDNA component (unfilled arrow). Insets at bottom show “empty” particles viewed close to an icosahedral five-fold axis where apical caps on opposite sides (front and back) of the particle nearly superimpose and appear as a ring in the center of the particle. Bar, 100 nm (main panel) and 50 nm (bottom insets).

Fig. 2

Shaded-surface representations of the ϕX17 4 reconstruction viewed along a two-fold axis (left) and along a five-fold axis (right). Bar, 15 nm.

Fig. 3

Upper portion of an equatorial slice of the reconstruction viewed along a two-fold axis. Arcs (at 10.0 and 15.5 nm radius) and tick marks (at 0.5-nm intervals) identify the radii corresponding to the spherical density projections in Fig. 4. Radial lines identify the positions of two-fold (2), three-fold (3), and five-fold (5) axes in the plane. Density at the largest radius in ϕX174 (16.8 nm) is indicated (arrowheads).

Fig. 4

Shaded-surface representations (first and third rows) and density projections (second and fourth rows) of the reconstruction truncated at successively lower radii (indicated in nanometers). The choice of a single threshold level for the surface representations at r = 12.5, 12.0, and 11.5 nm results in smooth spheres and gives a false impression of contiguous uniform density. The density projections at these radii give a more realistic rendering of the density distributions within the reconstruction.