Kehm E, Göksu M, Bayer S, Knopf C W
Deutsches Krebsforschungszentrum, Genomforschung und Bioinformatik, Heidelberg, Deutschland.
Intervirology. 1998;41(2-3):110-9. doi: 10.1159/000024922.
Herpes simplex virus type 1 DNase (HSV-1 DNase) was expressed in insect cells by recombinant baculovirus (NPVUL12) and purified by a combination of anionic exchanger chromatography and gel filtration. Two polypeptides of 85 and 75 kD, whose ratio varied during purification, were induced 24 h after infection. The 75-kD protein was isolated and shown to possess catalytic activity. Gel filtration analysis indicated that the active form of the enzyme at an ionic strength of I = 0.3 is a dimeric protein with an apparent molecular weight of 130,000. The recombinant enzyme exhibited the overall characteristics of the native enzyme such as 5'-3' exonuclease and endonuclease activities with a preferred degradation of DNA. In the absence of extraneously added Mg2+, the enzyme was capable of removing mononucleotides from 5'-end-labeled DNA, but not from RNA and 3'-end-labeled DNA. The peculiar mechanism of double-strand DNA degradation suggests a specific role of HSV-1 DNase in DNA recombination processes during viral replication.
单纯疱疹病毒1型脱氧核糖核酸酶（HSV-1 DNase）通过重组杆状病毒（NPVUL12）在昆虫细胞中表达，并通过阴离子交换色谱和凝胶过滤相结合的方法进行纯化。感染后24小时诱导出两种分子量分别为85kD和75kD的多肽，其比例在纯化过程中有所变化。分离出75kD的蛋白质并证明其具有催化活性。凝胶过滤分析表明，在离子强度I = 0.3时，该酶的活性形式是一种表观分子量为130,000的二聚体蛋白。重组酶表现出天然酶的总体特征，如5'-外切核酸酶和内切核酸酶活性，且优先降解DNA。在没有额外添加Mg2+的情况下，该酶能够从5'-末端标记的DNA中去除单核苷酸，但不能从RNA和3'-末端标记的DNA中去除。双链DNA降解的特殊机制表明HSV-1 DNase在病毒复制过程中的DNA重组过程中具有特定作用。
