Biology Reference
In-Depth Information
obtained either with the expression of a complete ORF2, a truncated
form of ORF2 lacking the first 70 amino acids, or after replacing
these 70 amino acids with the green fluorescent protein (GFP).
Recombinant HAstV VLPs will provide a virtually unlimited supply of
highly purified viral capsids with many immediate applications, such
as the characterization of antigenic properties of the mature capsid
proteins and epitope mapping studies. Additionally, VLPs may also be
used to determine the HAstV atomic structure using X-ray crystallo-
graphic techniques. Another potential application of the expressed
recombinant VLPs is to study cell-virus interactions in binding assays,
which may allow the identification of the cellular receptor involved in
viral infection.
Interestingly, during the formation of HAstV VLPs in insect cells,
a smaller second type of structure consisting of 16 nm ring-like units
was observed, mostly after disassembling the 38 nm VLPs through
the addition of EDTA.
27
As previously described for Norwalk virus as
well as for other T
3 RNA plant viruses, these 16 nm VLPs could
correspond to structures that are formed when 60 units of the capsid
protein assemble into a structure with T
=
1 symmetry. Depending on
factors such as trypsin digestion, the presence and size of the RNA
genome, the presence of divalent cations, the ionic strength and the
pH, capsid proteins of these viruses are able to self-assemble
in vitro
into T
=
=
3 or smaller T
=
1 structures.
44,45
Nonstructural Proteins
The nonstructural proteins of the virus are translated from the genomic
viral RNA as two polyproteins, one of which contains only ORF1a
(nsP1a: 101 kDa) and the other that includes ORF1a/1b (nsP1a1b:
160 kDa) and is translated via a -1 ribosomal frame-shifting event
between ORF1a and ORF1b.
11
Both proteins are proteolytically
processed, giving rise to a variety of proteins. While nsP1b (59 kDa) cor-
responds to the viral RNA-dependent RNA polymerase — which can be
aligned with other polymerases of the Koonin's supergroup I together
with picornaviruses, caliciviruses and certain plant viruses — little is
known about the role of most of the mature nsP1a products (Fig. 4).
The protease motif has features consistent with chymotrypsin-like
