[Kinase domain analysis of MDV-1 CVI988/Rispens UL13 and preferred codon fragments expression in Escherichia coli].
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 and antigenicity of UL13 in Escherichia coli was analyzed by online service GENEART (www.gcua.de)and DNAstar software respectively. 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.


CONCLUSIONS
The 152-297 residue is kinase catalytic center of MDV-1 UL13; UL13 protein expressed in vitro induced specific antibodies against UL13.
Zhang
Chenfei
C
Key Lab of Jiangsu Preventive Veterinary Medicine, Yangzhou University, Yangzhou 225009, China.
Qin
Aijian
A
Deng
Xufang
X
Su
Yuwen
Y
Xue
Min
M
Yin
Tianyan
T
Wang
Pingping
P
D000595:Amino Acid Sequence; D000818:Animals; D000914:Antibodies, Viral / Q000097:blood; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism; D015870:Gene Expression; D008381:Herpesvirus 2, Gallid / Q000737:chemistry / Q000201:enzymology / Q000235:genetics / Q000276:immunology*; D008380:Marek Disease / Q000276:immunology* / Q000821:virology; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D011494:Protein Kinases / Q000737:chemistry / Q000235:genetics / Q000276:immunology*; D017434:Protein Structure, Tertiary; D016415:Sequence Alignment; D014764:Viral Proteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology*
D000914:Antibodies, Viral; D014764:Viral Proteins; D011494:Protein Kinases