Kinase domain analysis of MDV-1 CVI988/Rispens UL13 and preferred codon fragments expression in Escherichia coli

Abstract:

Objective: To find catalytic center of MDV-1 UL13 and express it in vitro to investigate the function of UL13 kinase. Methods: UL13 gene was amplified by polymerase chain reaction (PCR) from MDV-1 CVI988/Rispens strain. The codon bias of UL13 in Escherichia coli was analyzed by online service GENEART (www.gcua.de）and DNAstar software. Then the UL13 truncated fragments were expressed in Escherichia coli, and mice were immunized with the expressed Glutathione S Transferase fusion protein. The conserved domain was analyzed with protein blast and Cn3D 4.1 online software of National Center for Biotechnology Information. Results: UL13 gene was successfully amplified. The sequence analysis suggests that 259-400 and 431-513 amino acid residues are low abundance for rare codon and strong antigenicity in UL13. Result of conserved domain analysis demonstrated that 152-297 residue iskinase catalytic center of UL13. However, conserved glycin in kinase subdomain Ⅶ for most protein kinase was replaced by serine in UL13 and proline in kinase subdomain Ⅷ replaced by cysteine. The serum from mice immunized with truncated fragment, 259-400 amino acids, could react with recombinant UL13 protein expressed in insect cells in immunofluorenscence assay. Conclusion: The 152-297 residue is kinase catalytic center of MDV-1 UL13; UL13 protein expressed in vitro induced specific antibodies against UL13.