Functional and Structural Study of Singapore Grouper Iridovirus ORF086R

Abstract

Singpaore grouper iridovirus (SGIV) is a major pathogen that causes significant economic losses in marine farms. The virus contains a dsDNA genome of about 140kb predicted to encode 162 open reading frames (ORFs). Our previous proteomics and transcriptomics study had identified ORF86R as an immediately early (IE) gene. We hypothesize that ORF86 may play an important role in virus infection and replication. Both full length and truncated ORF86R were cloned and expressed in E. coli to get soluble and stable protein for raising antibody and screening crystallization. Antisense morphlino oligonucleotide (asMOs) was used to knock down ORF86R expression in grouper embryonic cells. TCID50 and electron microscope were carried out to examine the viral infectivity and replication. We observed that TCID50 was reduced but viral morphology was not affected after the gene knockdown. Circular dichroism (CD) and dynamic light scattering (DLS) also showed that the protein is likely to be folded. The protein is mainly composed of ?-sheet structures by secondary structure prediction. Ongoing experiments are focused on identification of ORF86R?s binding partners and optimization of its crystallization conditions, which may help us understand how this viral IE gene interacts with host factors and facilitates its replication.