Viral envelope proteins have been proposed to play significant roles in
virus infection and assembly. In this study, an envelope
protein gene, 53R, was cloned and characterized from Rana grylio virus (RGV), a member of the family Iridoviridae. Database searches found its homologues in all sequenced iridoviruses, and sequence alignment revealed several conserved structural features shared by virus capsid or envelope
proteins: a myristoylation site, two predicted transmembrane domains and two invariant
cysteine residues. Subsequently, RT-PCR and Western blot detection revealed that the transcripts encoding RGV 53R and the
protein itself appeared late during
infection of fathead minnow cells and that their appearance was blocked by
viral DNA replication inhibitor, indicating that RGV 53R is a late expression gene. Moreover, immunofluorescence localization found an association of 53R with virus factories in RGV-infected cells, and this association was further confirmed by expressing a 53R-GFP fusion
protein in pEGFP-N3/53R-transfected cells. Furthermore,
detergent extraction and Western blot detection confirmed that RGV 53R was associated with virion membrane. Therefore, the current data suggest that RGV 53R is a novel
viral envelope protein and that it may play an important role in virus assembly. This is thought to be the first report on a
viral envelope protein that is conserved in all sequenced iridoviruses.