Expression and characterization of UL16 gene from duck enteritis virus

Qin He,Qiao Yang,Dekang Zhu,Zhengli Chen,Mingshu Wang,Anchun Cheng,Jun Xiang,Xiaoyue Chen,Qihui Luo,Renyong Jia,Yi Zhou

doi:10.1186/1743-422x-8-413

Abstract

BackgroundPrevious studies have indicated that the UL16 protein and its homologs from herpesvirus were conserved and played similar roles in viral DNA packaging, virion assembly, budding, and egress. However, there was no report on the UL16 gene product of duck enteritis virus (DEV). In this study, we analyzed the amino acid sequence of UL16 using bioinformatics tools and expressed in Escherichia coli Rosetta (DE3) induced by isopropy1-β-D-thiogalactopyranoside (IPTG). The recombinant protein was produced, purified using a Ni-NTA column and used to generate the polyclonal antibody against UL16. The intracellular distribution of the DEV UL16 product was carried out using indirect immunofluorescence assay.ResultsIn our study, UL16 gene of DEV was composed of 1089 nucleotides, which encoded 362 amino acids. Multiple sequence alignment suggested that the UL16 gene was highly conserved in herpesvirus family. The UL16 gene was cloned into a pET prokaryotic expression vector and transformed into Escherichia coli Rossetta (DE3) induced by IPTG. A 60kDa fusion protein band corresponding to the predicted size was produced on the SDS-PAGE, purified using a Ni-NTA column. Anti-UL16 polyclonal sera was prepared by immunizing rabbits, and reacted with a band in the IPTG induced cell lysates with an apparent molecular mass of 60 kDa. In vivo expression of the UL16 protein in DEV infected duck embryo fibroblast cells (DEFs) was localized mostly around perinuclear cytoplasmic area and in cytosol using indirect immunofluorescence assay.ConclusionsThe UL16 gene of DEV was successfully cloned, expressed and detected in DEV infected DEFs for the first time. The UL16 protein localized mostly around perinuclear cytoplasmic area and in cytosol in DEV infected DEFs. DEV UL16 shared high similarity with UL16 family members, indicating that DEV UL16 many has similar function with its homologs. All these results may provide some insight for further research about full characterizations and functions of the DEV UL16.

Highlights

Previous studies have indicated that the UL16 protein and its homologs from herpesvirus were conserved and played similar roles in viral DNA packaging, virion assembly, budding, and egress
Computer analysis showed that the UL16 amino acid sequence contains 16 possible sites for phosphorylation
Five casein kinase II, one cAMP- and cGMP- dependent protein kinase, five protein kinase C phosphorylation sites and four potential N-linked myristoylation sites were present along the amino acid sequence

Summary

Introduction

Previous studies have indicated that the UL16 protein and its homologs from herpesvirus were conserved and played similar roles in viral DNA packaging, virion assembly, budding, and egress. There was no report on the UL16 gene product of duck enteritis virus (DEV). Duck viral enteritis (DVE), an acute and contagious disease, is highly lethal in all ages of birds from the order Anseriformes (ducks, geese, and swans). This disease is characterized by vascular lesions and tissue hemorrhage, as well as gastrointestinal, lymphatic, and nervous impairments [1,2,3]. Duck enteritis virus (DEV) is the causative agent for DVE and was first recorded in Holland in 1923 [4], more outbreaks were reported in the. A lot of DEV genes have been identified and reported, such as glycoprotein B gene, glycoprotein E gene, thymidylate kinase gene, dUTPase pyrophosphatase gene et al [10,11,12]

Methods

Results

Conclusion