传染性造血器官坏死病毒-Sn1203株的基因型及糖蛋白的生物信息学分析

The genotype of infectious haematopoietic necrosis virus isolate IHNV-Sn1203 and the bioinformatics analysis of its glycoprotein

利用鲤（Cyprinuscarpio）上皮细胞（epitheliaomapapulosumcyprini，EPC）培养传染性造血器官坏死病毒．Sn1203分离株（IHNV-Sn1203），根据GenBank中IHNVG蛋白基因开放阅读框（openreadingflame，ORF）的序列设计引物（GenBank序列编号AB288207），采用RT-PCR的方法克隆得到IHNV-Sn1203株G蛋白全长ORF，克隆至表达载体pET27b（＋）中，构建了pET27-G重组质粒，并进行了测序分析。生物信息学分析结果显示，IHNV-Sn1203株G蛋白基因序列长度为1527bp，与韩国株具有最高的核酸同源性（96．86％）和氨基酸同源性（97．05％）。该基因编码508个氨基酸残基，推导分子量约为56．55kD，等电点为6．15；氨基酸序列分析表明，G蛋白富含丝氨酸、苏氨酸和酪氨酸，存在28个潜在的磷酸化位点；存在4个潜在的N-糖基化位点和7个潜在的O-糖基化位点；G蛋白N端含有20个氨基酸的信号肽；亲水性大于输水性；位于483～508位氨基酸存在一跨膜区；抗原表位预测显示抗原性良好；系统进化树分析显示，IHNV-Snl203株与日本株和韩国株聚为一簇，都属于JRt基因型。

The glycoprotein （G） is one of the structural proteins on the surface of infectious haematopoietic necrosis virus （IHNV） and has important functions. In this study, the open reading frame of the glycoprotein gene was amplified by reverse transcriptase-PCR （RT-PCR） from isolate IHNV-Snl203 and cloned into the pET27b（＋） vector. The structure and characteristics of the glycoprotein gene were analyzed using bioinformatics software. The results showed that the G gene was 1 527 bp, encoding a 508 amino acid protein, Isolate IHNV-Snl203 shared closer identity with strains isolated from South Korea, and the homology of at nucleotide and amino acid level of the G gene were 96.68% and 97.05%, respectively. The molecular weight was 56.55 kD and the isoelectric point was 6.15. The glycoprotein was rich inserine, threonine and tyrosine, and contained 28 potential phosphorylation sites. According to the protein structure prediction, the glycoprotein may have four N-glycosylationsites and seven O-glycosylation sites. The glycoprotein is hydrophilic, with an N-terminal signal peptide and a transmembrane region from amino acid 483 to 508. The glycoprotein was esti- mated to be highly antigenic. Phylogenetic analysis showed that isolate IHNV-Snl203 was in the same branch as iso- lates from Japan and South Korea, belonging to the JRt genotype. This study established a foundation for research into the ~enetic background, oathogenesis, and molecular enidemiology of IHNV.