particle as it is scanned. Deflections of a laser beam focused on the tip during scanning are recorded via a photodiode, allowing construction of an image of the particle surface. When the force applied via the tip is increased, the degree of deformation of the particle is measured as a function of force, as illustrated in the center. As the force increases, the virus particle eventually breaks, leading to a sudden drop in resistance (bottom). This force is often called the breaking force. Courtesy of G. Nemerow, The Scripps Research Institute. (B) Shown in the right two panels are atomic force microscopy images of a particle of human adenovirus type 5 with short fibers derived from type 35 before and after application of sufficient force to induce mechanical failure. The degree of resolution is low, but sufficient to determine the orientation of these icosahedral particles on the surface. To characterize mechanical failure in more detail, the responses of individual particles were scored on the basis of whether an edge, a facet, or a vertex was probed, revealing three distinct patterns (left three panels). This approach established that the vertices are the weakest points, exhibiting the lowest spring constant and breaking force. Adapted from Snijder J et al. 2013. J Virol 87:2756–2766.
Stabilization and Destabilization of Virus Particles
In all but the simplest T = 1 icosahedral capsids, there is some degree of mismatch between the pentons at the 12 vertices and the hexons that surround them. The frequent reinforcement of these associations at the vertices by especially extensive interactions among structural proteins, as in polioviruses (Fig. 4.13C), or by specialized “cement” proteins, as in human adenoviruses (Fig. 4.16B), is not therefore surprising. Nevertheless, the vertices of icosahedral capsids that have been examined by nanoindentation are the points most susceptible to breakage (Fig. 4.30B). Indeed, human adenovirus pentons are the first components to dissociate under mild pressure in vitro as well as during cell entry. Furthermore, binding of integrin, the coreceptor for entry of these viruses (Chapter 5), to pentons further weakens the capsid.
Among the minor capsid proteins of herpes simplex virus type 1 are two that bind both pentons and neighboring hexons. As might be anticipated, capsids that lack either of these proteins exhibit decreased stiffness. In some cases, the presence of the viral genome substantially increases capsid stability. For example, comparison of the mechanical properties of full and empty capsids of the (+) strand insect virus triatoma virus by nanoindentation identified pH-dependent changes: at neutral pH, as in infected cells, mature virus particles were some threefold stiffer than those without genomes and more resistant to deformation. However, these properties were reversed at a more alkaline pH, like that of the hindgut of the insect host, where the virus encounters host cells. Binding of the single-stranded DNA genome of the parvovirus minute virus of mice also increases the stiffness of virus particles, and concomitantly their resistance to thermal inactivation.
A variety of viruses are assembled as immature forms that are converted to infectious particles upon proteolytic processing of structural protein precursors. It is now clearly established that such proteolytic cleavages are accompanied by mechanical alterations that increase internal pressure to facilitate DNA ejection (e.g., herpesviruses and many bacteriophages) or decrease the mechanical strength of virus particles to facilitate disassembly (e.g., human adenoviruses) or entry into the host cell (e.g., human immunodeficiency virus). Such mechanical transformations are considered in Chapters 5 and 13.
Perspectives
Virus particles are among the most elegant and visually pleasing structures found in nature, as illustrated by the images presented in this chapter. Now that many structures of particles or their components have been examined, we can appreciate the surprisingly diverse architectures they exhibit. Nevertheless, the simple principles of their construction proposed more than 60 years ago remain pertinent: with few exceptions, the capsid shells that encase and protect nucleic acid genomes are built from a small number of proteins arranged with helical or icosahedral symmetry. This feature is characteristic of even some of the largest viruses yet described, indicating that not only genetic economy but also optimized and regular interactions among structural units dictate virus architecture.
The detailed views of nonenveloped virus particles provided by X-ray crystallography emphasize just how well these protein shells provide protection of the genome during passage from one host cell or organism to another. They have also identified several mechanisms by which identical or nonidentical subunits can interact to form icosahedrally symmetric structures, and protein-protein interactions that stabilize larger virus particles with icosahedral symmetry. More-elaborate virus particles, which may contain additional protein layers, a lipid envelope carrying viral proteins, and enzymes or other proteins necessary to initiate the infectious cycle, pose greater challenges to the structural biologist. Indeed, for many years we possessed only schematic views of these structures, deduced from negative-contrast electron microscopy and biochemical or genetic methods of analysis. In previous editions, we noted the power and promise of continuing refinements in methods of cryo-EM (or cryo-electron tomography), image reconstruction, and difference imaging. These techniques have attained atomic- or near-atomic-level resolution, providing remarkable views of large viruses with multiple components, viral envelopes, and, in some cases, the organization of genomes within particles. The structural descriptions of ever-increasing numbers of viruses representing diverse families have also allowed unique insights into evolutionary relationships among seemingly disparate viruses or viral proteins.
These extraordinary advances notwithstanding, important challenges remain, most obviously the visualization of structures that do not exhibit simple symmetry (or are not constructed from components that do). These structures include many genomes and the particles of some large viruses (e.g., poxviruses). The giant viruses, such as mimiviruses and pithoviruses, some with particles so large that they can be seen by light microscopy, also pose new technical challenges and intimate that unanticipated structural principles remain to be elucidated.
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