THE CHARACTERIZATION OF THE EBV ALKALINE DEOXYRIBONUCLEASE CLONED AND EXPRESSED IN ESCHERICHIA-COLI
THE CHARACTERIZATION OF THE EBV ALKALINE DEOXYRIBONUCLEASE CLONED AND EXPRESSED IN ESCHERICHIA-COLI
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DOI：
10.1093/nar/17.19.7609
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发表时间：
1989-10-11
影响因子：
14.9
通讯作者：
LITTLER, E
中科院分区：
文献类型：
--
作者：
BAYLIS, SA;PURIFOY, DJM;LITTLER, E
核酸同源性研究表明，EB病毒（EBV）的BGLF 5开放阅读框架编码一种碱性脱氧核糖核酸酶（DNase），与单纯疱疹病毒的DNase有一定的同源性。本文报道了BGLF 5开放阅读框在大肠杆菌中的表达。大肠杆菌中表达的一种新的碱性DNA酶的活性高水平诱导细胞。这种碱性DNA酶已被纯化至表观均一性作为单一蛋白质种类。本文首次报道了疱疹病毒DNA酶在原核系统中的表达及纯化。它具有典型疱疹病毒碱性核酸外切酶的生物化学特征，表现出高pH最适值，对Mg 2+的绝对需求，以及对高盐浓度和多胺的敏感性。该酶活性可被鼻咽癌患者血清中和，并与鼻咽癌患者血清发生反应。因此，本文所述的原核表达系统为生物化学和血清流行病学分析提供了经济有效的EBV DNA酶来源。
Studies of nucleic acid homology suggest the BGLF5 open reading frame of Epstein-Barr virus (EBV) encodes an alkaline deoxyribonuclease (DNase) sharing some homology with that of herpes simplex virus. We report here the expression of the BGLF5 open reading frame in E. coli and the expression of high levels of a novel alkaline DNase activity in induced cells. This alkaline DNase has been purified to apparent homogeneity as a single protein species. This is the first report of the expression of a herpesvirus coded DNase in a prokaryotic system and of the purification fo the EBV DNase to demonstrable purity. It has the biochemical characterstics of a typical herpesvirus alakaline exonuclease showing a high pH optimum, an absolute requirement for Mg2+ for activity and sensitivity to high salt concentrations and polyamines. The enzyme activity was neutralized by sera from patients with nasopharyngeal carcinoma and was reactive with these sera in Western blot analysis. Thus the prokaryotic expression system described here provides an economical and efficient source of the EBV DNase for biochemical and seroepidemiological analysis.