Human enteric Caliciviridae: the complete genome sequence and expression of virus-like particles from a genetic group II small round structured virus

Abstract

Comparisons of the RNA polymerase and capsid sequences of small round structured viruses (SRSVs) have recently shown these are genetically diverse viruses which fall into two distinct groups. The genomes of two group I viruses, Southampton and Norwalk viruses have been characterized; however, similar data for the genetic group II SRSVs have not been available until now. We report here the complete genome sequence of a recent group II SRSV, Lordsdale virus. The Lordsdale virus genome is 7555 nt in length and has a similar organization to the group I SRSVs. The large ORF in the 5' half of the genome (5100 nt) is shorter than the group I SRSV ORF1 (5367 nt), but has the characteristic 2C helicase, 3C protease and 3D RNA polymerase enzyme motifs. ORF2, encoding the structural protein is of a similar size to the group I viruses but the small 3'-terminal ORF is significantly larger in group II. A highly conserved sequence of 28 nt was identified at the start of Lordsdale virus ORF1 and repeated at the start of ORF2. These conserved motifs are typical of the animal caliciviruses. Comparison of the 150 N-terminal amino acids in the ORF1 protein revealed little identity between the two SRSV genetic groups, reflecting the shorter ORF1 in the group II virus. Recombinant baculoviruses containing ORF2 and ORF3 sequences were constructed and used to express large quantities of the group II Lordsdale virus structural protein. The capsid protein formed virus-like particles by self assembly which resembled ' empty' SRSVs.

Efficient assembly and release of SARS coronavirus‐like particles by a heterologous expression system

“…In our study, the expression of the S protein was very high and the incorporation of S glycoprotein onto the surface of the VLPs makes these particles not only a powerful instrument for studies of the assembly and budding of SARS-CoV particles but also as a candidate vaccine and a tool for SARS diagnosis. Self-assembly of viral capsid proteins into VLPs for both RNA and DNA viruses has been reported and VLPs produced by this system usually retain the immunogenic and physiochemical properties of the native virions [27][28][29][30][31][32].…”

Genetic classification of "Sapporo-like viruses"

“…In addition, in all published SLV strains except a human strain, London/92 and a porcine strain, PEC/Cowden [5,15,18,27,29,40], an additional ORF has been predicted in +1 frame, overlapping the N terminus of the capsid gene (capsid overlap). To date, the sequence of a ∼3-kb region extending from the RNA polymerase gene to the 3 poly(A) tail is available in GenBank for 9 NLV strains, of which 6 have had their entire genome sequenced: Norwalk/68 [17], Southampton/91 [22], Lordsdale/93 [9], Camberwell/94 [3,36], Chiba/87 (GenBank accession number No. AB042808) and Jena/80 [26].…”

Characterization of Monoclonal Antibodies Generated against Norwalk virus Gil Capsid Protein Expressed in Escherichia coli

“…The capsid protein of NV has an apparent molecular mass of 60 kDa. Expression of cDNAs that encode NV capsid proteins in insect cells infected with recombinant baculoviruses has been reported, demonstrating that all expressed recombinant proteins spontaneously assemble into empty virus-like particles (VLPs) (2,4,6,8,12,16). Since NV cannot be propagated in vitro, the availability of large amounts of recombinant NV (rNV) VLPs has facilitated the production of polyclonal antibodies as well as monoclonal antibodies (MAbs) useful to develop immunological detection systems and characterize anti- system to detect all types of NV (various clusters of NVs in two genogroups ) has not yet been established.…”