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of the RNA and production of a polyprotein ensues. The polyprotein is then processed by viral proteases
to form the viral polypeptides. The coat must then assemble, package RNA, and leave the cell. The
capsid-binding compounds' effect on propagation have been shown to occur at the attachment,
uncoating, and assembly steps (25,27–30). The relative effect on each of these steps appears to be
variable and has not yet been extensively characterized. The most universal effect appears to be on the
uncoating step.
During uncoating several different rhinovirus subparticles are observed. These are thought to be
intermediates in the uncoating process [32–33]. The fully infectious 149S particle becomes an 125S Aparticle,
which has lost VP4 but still maintains viral RNA. This A-particle is no longer infectious. The Aparticle
then releases RNA to become an 80S empty shell.
The formation of these types of particles can be induced by association of the virion with the cell
receptor or acidification [32,34]. In poliovirus, attachment to cells has been shown to lead to a particle
that has lost VP4 and has externalized the N-terminal region of VP1, which normally resides on the
virion interior.
There has been considerable debate about how the HRVs accomplish the transfer of RNA from inside
the virion into the cell cytosol. This step is crucial for productive uncoating. An important question
concerns the requirement for acidification of the endosome for HRVs to release their RNA. Evidence
that appeared to conflict was found in a number of studies using either entero- or rhinoviruses (35–39).
This question was later addressed by experiments that specifically separated entero- and rhinovirus
behavior [40]. These experiments showed that HRVs, unlike poliovirus, require a pH-lowering step for
productive infection. This pH lowering is likely to occur in the endosomal compartment. It should be
noted that HRV and enteroviruses have been classified historically based on their resistance to acid:
HRVs are acid-labile, while enteroviruses are stable in acid. Consequently, differences in behavior
between the rhino- and enteroviruses in an acidic environment within the cell are not surprising.
To replicate the changes in HRV that might be induced by acidification in the endosome, HRV14 was
acidified in the crystalline state and examined via x-ray diffraction. When compared to the native
HRV14, the acidified HRV14 capsid becomes disordered in three regions: the Ca 2+ ion on the five-fold
axis; a region of the β cylinder and the adjacent portion of VP4; and the GH loop [41]. The GH loop is
the region of structure that connects β-strands G and H and lies directly between the hydrophobic pocket
and the receptor binding site (Figure 4). It forms the roof of the VP1 hydrophobic pocket and the floor of
the canyon at the receptor binding site.
Mutants that are resistant to acid in vitro were isolated [41,42]. These mutants cluster about the GH loop
(Table 1, Figure 4), and typically would be thought of as mutants that stabilize protein structure (e.g.,
larger or more
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