锦鲤疱疹病毒ORF72基因克隆及结构功能分析

Cloning and Functional Analysis of ORF72 Gene in Koi Herpes Virus

于辽宁丹东采集感染锦鲤疱疹病毒的活鲤Cyprinus carpio样品,经DNA提取、PCR扩增、重组质粒构建步骤成功克隆锦鲤疱疹病毒ORF72基因,并利用生物信息学软件分析ORF72蛋白结构与功能,构建系统进化树。结果显示:ORF72基因开放阅读框共1113bp,编码蛋白由370个氨基酸组成,理论相对分子质量为40.5KD;ORF72蛋白主要由α螺旋、β折叠、无规则卷曲3种二级结构组成;N端35个氨基酸序列为信号肽序列。跨膜结构分析表明:ORF72蛋白无跨膜区,保守结构域预测表明ORF72含有一个保守结构域,抗原表位分析表明,抗原性较好。结构预测显示:ORF72氨基酸序列存在2个潜在的N-糖基化位点、1个潜在的O-糖基化位点、19个潜在的磷酸化位点。系统进化树分析发现,其与锦鲤疱疹病毒美国株、以色列株属同一分支。

The samples of common carp （Cyprinus carpio kio） infected with koi herpes virus were collected from Dandong area in Liaoning province, and ORF72 gene was successfully cloned by DNA extraction, PCR amplification, and plasmid recombination to analyze the structure and function ofORF72 gene and to construct a phylogenetic tree. The ORF72 was found to be composed of 1 113 nu- cleotides encoding a protein of 370 amino acids with a molecular weight of 40.5 KD. The ORF72 secondary structure was primarily comprised ofct helix, [3 strand, and random coil, and contained a conserved domain with N-terminal 35 residues as the signal peptides. Online analysis revealed that there was no transmembrane region in the ORF72 showing good antigenicity. The protein structure analysis indicated that there were two N-glycosylation sites, one O-glycosylation site and 19 phosphorylation sites. Phylogenetic analy- sis showed that ORF72 gene of KHV-DD was in the same branch with the USA and Israel strains.