Classical Swine Fever Virus Suri주의 gp44 유전자 클론닝과 염기서열 분석

Classical swine fever virus (CSFV) Suri strain is a variant derived from LOM vaccine virus, characterized with negative reaction in END (exaltation of Newcastle disease virus) test and with positive reaction in interference test of Western equine encephalitis virus. In this study, gp44 ($E^{rns}$) gene of Suri strain was amplified by the RT-PCR and the nucleotide and amino acid sequences were determined and analyzed to elucidate the molecular properties of Suri strain. The results obtained through the experiments were summarized as follows. The gp44 genes of ALD, Suri and LOM strains were amplified by RT-PCR using gp44-1S/AS and gp44-2 S/AS primer pairs. And all of strains showed the bands at 364 bp and 567 bp, respectively. As used gp44-1S and gp44-2AS primers, the 885 bp fragment, the whole gene of gp44, were specifically amplified in all of strains. The amplified cDNA of 885 bp was inserted into BamHI site of pAcGP67B baculovirus transfer vector and designated as pAcgp44 plasmid. As compared the 885 bp nucleotide and the amino acid sequences of gp44 of Suri strain with those of other viruses, ALD and GPE-, a vaccine strain in Japan, showed high homology ($97.2{\sim}98.5%$), and Brescia and Alfort, $91.9{\sim}94.6%$ homology. However, BVDV-NADL showed the lowest homology of $69.7{\sim}74.6%$. The phylogenic trees established on the basis of these results showed that ALD, GPE- and Suri had the highest relation, being classified in a same cluster, but BVDV-NADL maintained the long distance from the strains of CSFV. In hydrophobicity analysis of amino acid, all of CSFV except BVDV-NADL revealed the analogous patterns of hydrophobicity. As C2 and C3 conserved domains of amino acid sequences of gp44 protein of Suri strain were compared with those of other viruses, Suri strain showed the highest homology ($99.1{\sim}98.1%$) with ALD and GPE-, and comparably the lower homologies, with Alfort, Brescia and BVDV-NADL. C3 domain was more conserved than C2 domain. As compared the. structures of N-glycosylation sites of gp44 of Suri strain with the various strains, ALD and Brescia were exactly coincident with those of Suri strain in the locations and numbers. But GPE-, Alfort and BVDV-NADL showed the different structures. The locations and numbers of cysteine residues in gp44 were same in all CSFV except BVDV-NADL. All of six pestiviruses showed the same locations and numbers of the histidine residues in two RNase active sites located in gp44.