Immunogenicity of the capsid precursor and a nine-amino-acid site-directed mutant of the 3C protease of foot-and-mouth disease virus coexpressed by a recombinant goatpox virus

• Published in: Archives of virology 2014-07, Vol.159 (7), p.1715-1722
• Main Authors: Ma, Wenge, Wei, Jie, Wei, Yurong, Guo, Huiling, Jin, Yinghong, Xue, Ying, Wang, Yan, Yi, Zhong, Liu, Liya, Huang, Jiong, Wang, Lijian
• Format: Article
• Language: English
• Subjects: 3C Viral Proteases; Amino Acid Sequence; amino acids; Animals; Antibodies, Neutralizing - blood; Antibodies, Viral - blood; antigens; Antigens, Viral - genetics; Antigens, Viral - immunology; Biomedical and Life Sciences; Biomedicine; Capripoxvirus; Capripoxvirus - genetics; Capripoxvirus - metabolism; capsid; Capsid Proteins - genetics; Capsid Proteins - metabolism; Cysteine Endopeptidases - genetics; Cysteine Endopeptidases - metabolism; Electron microscopy; Female; Foot & mouth disease; Foot-and-mouth disease; Foot-and-mouth disease virus; Foot-and-Mouth Disease Virus - genetics; Foot-and-Mouth Disease Virus - metabolism; Gene Expression Regulation, Viral - physiology; genes; Genomes; Goatpox virus; immune response; Immunization; Immunogenicity; Infectious Diseases; Insertion; intergenic DNA; loci; Mammalian cells; Medical Microbiology; Mice; Mice, Inbred BALB C; Mutagenesis; Mutagenesis, Site-Directed; Mutants; Mutation; neutralizing antibodies; Original Article; Protective antigen; Proteinase; proteinases; proteins; Proteolysis; sheep; Site-directed mutagenesis; Theileria; Tissue polypeptide antigen; TPA; Vaccines, Synthetic - immunology; Viral Proteins - genetics; Viral Proteins - metabolism; Viral Vaccines - immunology; Virology; Virus-like particles; Viruses; VP1 protein; VP3 protein; Western blotting
• ISSN: 0304-8608; 1432-8798
• DOI: 10.1007/s00705-014-1984-8

The myristoylated capsid precursor mP1-2A of foot-and-mouth disease virus (FMDV), when expressed in mammalian cells and processed by the FMDV 3C protease, can self-assemble into virus-like particles (VLPs). In the present study, nine amino acids of the 3C protease were replaced by site-directed mutagenesis to create a mutant 3C protease, 9m3C. To coexpress mP1-2A and 9m3C and test the resulting proteolytic processing and VLP assembly, two recombinant goatpox viruses (rGTPVs) were constructed by the insertion of two coding regions, one for mP1-2A and the other for either 9m3C (rGTPV-mP1-2A-9m3C) or Theileria protective antigen (TPA) as a control (rGTPV-mP1-2A-TPA). The two exogenous genes were inserted into an intergenic region between loci gp_24 and gp_24.5 of the rGTPV genome. Western blotting of cells infected with rGTPV-mP1-2A-9m3C showed that proteins VP0, VP1, and VP3 from the mP1-2A processed by the 9m3C protease could be detected by polyclonal FMDV sera. As observed by electron microscopy, the infected cells produced VLPs with a diameter of about 25 ± 2 nm. Titers of neutralizing antibody against FMDV were significantly higher in mice inoculated with rGTPV-mP1-2A-9m3C, which expresses the 9m3C protease together with mP1-2A, than mice inoculated with the control rGTPV-mP1-2A-TPA, which does not express the protease. An ovine immunization test determined that sheep inoculated intramuscularly with rGTPV-mP1-2A-9m3C produced FMDV-specific neutralizing antibody, but its titers did not meet the requirement of the World Organization for Animal Health. The result indicates that further modifications of rGTPV-mP1-2A-9m3C are necessary to produce an effective vaccine.