title: Cysteine residues of the porcine reproductive and respiratory syndrome virus ORF5a protein are not essential for virus viability.
abstract: ORF5a protein was recently identified as a novel structural protein in porcine reproductive and respiratory syndrome virus (PRRSV). The ORF5a protein possesses two cysteines at positions 29 and 30 that are highly conserved among type 2 PRRSV. In this study, the significance of the ORF5a protein cysteine residues on virus replication was determined based on a type 2 PRRSV cDNA clone (pAJXM). Each cysteine was substituted by serine or glycine and the mutations were introduced into pAJXM. We found that the replacement of cysteine to glycine at position 30 was lethal for virus viability, but all serine mutant clones produced infectious progeny viruses. This data indicated that cysteine residues in the ORF5a protein were not essential for replication of type 2 PRRSV. The bimolecular fluorescence complementation (BiFC) and Co-immunoprecipitation (Co-IP) assay were used to study ORF5a protein interacted with other enveloped proteins. These results showed that ORF5a protein interacted non-covalently with itself and interacted with GP4 and 2b protein. The replacement of cysteine to glycine at position 30 affected the ORF5a protein interacted non-covalently with itself, which may account for the lethal phenotype of mutants carrying substitution of cysteine to glycine at position 30.
authors.affiliation: Department of Swine Infectious Diseases, Shanghai Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Shanghai 200241, PR China; Key Laboratory of Animal Diseases Diagnostic and Immunology, College of Veterinary Medicine, Nanjing Agricultural University, Ministry of Agriculture, Nanjing 210095, PR China.
authors.forename: Lichang
authors.initials: L
authors.lastname: Sun
authors.affiliation: Department of Swine Infectious Diseases, Shanghai Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Shanghai 200241, PR China.
authors.forename: Yan
authors.initials: Y
authors.lastname: Zhou
authors.affiliation: Department of Swine Infectious Diseases, Shanghai Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Shanghai 200241, PR China.
authors.forename: Runxia
authors.initials: R
authors.lastname: Liu
authors.affiliation: Department of Swine Infectious Diseases, Shanghai Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Shanghai 200241, PR China.
authors.forename: Yanhua
authors.initials: Y
authors.lastname: Li
authors.affiliation: Department of Swine Infectious Diseases, Shanghai Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Shanghai 200241, PR China.
authors.forename: Fei
authors.initials: F
authors.lastname: Gao
authors.affiliation: Department of Swine Infectious Diseases, Shanghai Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Shanghai 200241, PR China.
authors.forename: Xiaomin
authors.initials: X
authors.lastname: Wang
authors.affiliation: Key Laboratory of Animal Diseases Diagnostic and Immunology, College of Veterinary Medicine, Nanjing Agricultural University, Ministry of Agriculture, Nanjing 210095, PR China.
authors.forename: Hongjie
authors.initials: H
authors.lastname: Fan
authors.affiliation: Department of Swine Infectious Diseases, Shanghai Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Shanghai 200241, PR China.
authors.forename: Shishan
authors.initials: S
authors.lastname: Yuan
authors.affiliation: Department of Swine Infectious Diseases, Shanghai Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Shanghai 200241, PR China; College of Animal Science and Technology, Guangxi University, Nanning 530005, PR China. Electronic address: zuzhangwei@163.com.
authors.forename: Zuzhang
authors.initials: Z
authors.lastname: Wei
authors.affiliation: Department of Swine Infectious Diseases, Shanghai Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Shanghai 200241, PR China. Electronic address: gztong@shvri.ac.cn.
authors.forename: Guangzhi
authors.initials: G
authors.lastname: Tong
mesh_terms: D019943:Amino Acid Substitution; D000818:Animals; D002460:Cell Line; D017124:Conserved Sequence; D003545:Cysteine / Q000235:genetics / Q000378:metabolism*; D004252:DNA Mutational Analysis; D050296:Microbial Viability; D019316:Porcine respiratory and reproductive syndrome virus / Q000235:genetics / Q000502:physiology*; D013552:Swine; D014759:Viral Envelope Proteins / Q000235:genetics / Q000378:metabolism*; D014779:Virus Replication
chemical_list: D014759:Viral Envelope Proteins; C114789:glycoprotein 5, PRRSV; D003545:Cysteine
keywords: Cysteine; ORF5a; PRRSV; Replication