Alphaviruses are small 70 nm viruses that have 240 copies each of three structural proteins, E1, E2 and capsid (C) assembled in a 1:1:1 stoichiometry. These three proteins create two nested T = 4 icosahedral shells that sandwich a host derived lipid bilayer. The outer protein shell is composed of E1 and E2 heterodimers that assemble into aggregates of three producing the three pronged spike which protrudes from the virus surface. The inner protein shell is made of only C, encapsulating the 49S RNA. This inner protein shell is the shape determining component of the virus. The organization of the E1/E2 complexes on the cell membrane likely exist in a two-dimensional 6-fold symmetry sheet prior to capsid envelopment. When the membrane glycoproteins begin the process of encapsulation, the nucleocapsid recruits the E1-E2 trimers into the developing outer shell by specifically binding the E2 endodomains. Through its repeated 240 interactions between a hydrophobic cleft on C and the E2 endodomain this process organizes the glycoproteins into the 6-fold and requisite 5-fold rotational arrays necessary to form a three-dimensional icosahedral structure. Mutations which disrupt this process result in structurally misshaped particles. The resulting virus particle has 80 spikes that are primarily made of E2 (colored blue in Figure 1 pH 7.0) with a protein skirt that is primarily E1 (colored green in Figure 1 pH 7.0) which covers the incorporated membrane. The two protein shells with their significant level of lateral and transmembrane interactions result in a very rigid and precisely organized particle that is unlike that of other membrane containing viruses. It has been shown recently, that the particles of mammalian and insect grown SVHR are not structurally identical. The cellular response to infection by insect and mammalian derived virus has also been shown to be different. Particles produced from insect cells are more compact, lacking some RNA intercalation in the capsid protein shell seen in mammalian grown virus. The thickness of the membrane of virus produced from mosquito and mammalian cells does not solely account for the difference in the structure of the virus particles; however the outer protein shell seems to be extended in the mammalian grown virus suggesting that the protein organization between the two particles may be in slightly different functional conformations.
The highly organized structure of the alphaviruses with the many protein-protein interactions stabilizing the structure and the membrane bilayer occluded by the outer protein shell have important implications for the process of entry of the viral genome into host cells. Whereas many membrane containing viruses such as influenza can be described as a membranous structure with embedded proteins the alphaviruses are protein shells with an associated membrane. This protein shell must be compromised if the virus is to transfer its RNA to the cell cytoplasm. The structure of the virus protein shell may be critical for the mechanism of genome delivery to work properly. It is hypothesized that the E1 protein in the mature alphavirus exists in a metastable configuration poised to use the energy stored during virus assembly for the entry process upon encountering an appropriate trigger. The energy stored for this conformational change is the result of folding the E1 protein though a series of disulfide bonded intermediates as the protein is compacted into the metastable structure during the assembly of the spike heterotrimer. The metastable nature of the spike complex is revealed when the E1 protein is released from the mature virus using detergent because the protein reorganizes into several disulfide bridged configurations which can be separated on non-denaturing gels.