Virus maturation: dynamics and mechanism of a stabilizing structural transition that leads to infectivity

Abstract

For many viruses, the final stage of assembly involves structural transitions that convert an innocuous precursor particle into an infectious agent. This process -- maturation -- is controlled by proteases that trigger large-scale conformational changes. In this context, protease inhibitor antiviral drugs act by blocking maturation. Recent work has succeeded in determining the folds of representative examples of the five major proteins -- major capsid protein, scaffolding protein, portal, protease and accessory protein -- that are typically involved in capsid assembly. These data provide a framework for detailed mechanistic investigations and elucidation of mutations that affect assembly in various ways. The nature of the conformational change has been elucidated: it entails rigid-body rotations and translations of the arrayed subunits that transfer the interactions between them to different molecular surfaces, accompanied by refolding and redeployment of local motifs. Moreover, it has been possible to visualize maturation at the submolecular level in movies based on time-resolved cryo-electron microscopy.

Figure 1

Generic assembly pathway for the heads of tailed bacteriophages.

Figure 2

Structural genomics of capsid assembly. Diagrams of the procapsid and mature head (capsid plus internally coiled DNA) are shown at center (from Figure 1). Ribbon diagrams are shown for the φ29 scaffold (PDB code 1N04, [13^••]) and portal/connector (PDB code 1IJG, [9]), the HSV2 protease (PDB code IAT3, [61]), the λ accessory protein (PDB code 1TCZ, [26]) and the HK97 MCP (PDB code 1OHG, [19]). It remains to be seen whether system-to-system variation is accomplished by embellishment of the same basic fold (e.g. by adding additional domains, as with portals, whose molecular masses vary widely [12^••]) or by substituting proteins with entirely different folds. We conjecture that accessory proteins and scaffolding proteins are the likeliest candidates for variations of the latter kind.

Figure 3

Maturation pathway of the HK97 capsid, visualized in surface renderings (outer surface, upper rows; inner surface, lower rows) at ~14 Å resolution. All images are from cryo-EM reconstructions, except Head II, which is a resolution-limited rendition of the crystal structure [19]. The capsid has icosahedral geometry (T=7 laevo) and is viewed along a twofold axis of symmetry. Prohead I is composed of 420 copies of gp5. In Prohead II, the N-terminal Δ domains have been removed from the inner surface. In acid-induced maturation in vitro, the first transition state, EI-I (not shown), is about 10% bigger than Prohead II [29] and very similar to EI-II [33^••] (shown). The main difference between EI-I and EI-II is that EI-II has some covalent cross-links [31^•]. The next structural state is a thin-walled spherical particle called the ‘balloon’. Balloons vary in their extents of cross-linking, as in EI-III (variable partial cross-linking) and EI-IV (almost complete cross-linking). The balloon structure is very similar to that of the end-state Head, except for the positions of its pentons, which move ~30 Å outwards in the final transition. Bar = 100 Å .

Figure 4

Movements of and VP26 binding by the protrusion domains of HSV1. (a) Maturation of the 1250 Å diameter HSV1 procapsid (left), whose shell contains VP5 (the MCP, blue) and the triplex proteins, VP19c and VP23 (green). If the portal protein, UL6, is available, it occupies one vertex. The scaffolding protein, preUL26.5, and the protease–scaffold fusion, preUL26, form an inner shell that is not shown. The matured capsid (middle) exposes binding sites around the tips of the VP5 hexamers that bind six copies of VP26 (orange) — right. VP26 does not bind to penton VP5. The differing conformations of penton and hexon VP5 are denoted by differing shades of blue in all three panels. The binding site for VP26 on hexons involves two adjacent subunits of VP5: inappropriate juxtaposition of these subunits offers an explanation for the failure of VP26 to bind to procapsid hexons or pentons (either state). The left and middle images were adapted from [44^••], and the right from [62]. (b) The crystal structure of the protrusion domain (PDB code 1NO7, [40^••]) has been fitted into the P-hexons (i.e. the peripentonal hexons) of the procapsid (left) and the mature capsid (right). The initial fitting was done by hand and then refined with an automated program (JB Heymann, unpublished). In the precursor state, there is essentially no contact between adjacent protrusion domains. In maturation, they swivel about hinges at the top of the underlying floor domain to make extensive nearest-neighbor contacts in a now highly sixfold symmetric hexon protrusion. The cryo-EM envelope of the capsid is blue. Ribbon diagrams of alternating protrusion domains around a hexon are pink and yellow.