Lin S F, Hsu T Y, Liu M Y, Lin L S, Yang H L, Chen J Y, Yang C S
Graduate Institute of Microbiology, College of Medicine, National Taiwan University, Taipei.
Virology. 1995 Apr 20;208(2):712-22. doi: 10.1006/viro.1995.1203.
Bacterially expressed Epstein-Barr virus (EBV) DNase was purified to 98% purity and used as the source for characterization of the enzyme activities. Complete digestion of DNA by EBV DNase yielded 5'-monophosphate nucleosides as the final products. During the logarithmic phase of the reaction, EBV DNase acted processively on dsDNA but distributively on ssDNA. Both 5' to 3' and 3' to 5' exonuclease activities were present, although the former was shown to be 10-fold stronger. No significant discrepancy was seen in the liberation of end-labeled nucleotides by DNase when substrates with 5'-protruding, blunt, or 3'-protruding ends were used. EBV DNase was demonstrated also to have an endonuclease activity using supercoiled plasmid DNA as substrate. Two preferential dsDNA cleavage sites were mapped on pBS-TR, a pBlueScript vector containing one copy of the EBV terminal repeat; both are in vector sequences. Finally, an N-terminally truncated EBV major DNA binding protein, but not EA-D, was shown to inhibit EBV DNase activity. This inhibitory effect may due to direct protein-protein interactions between EBV DNase and the major DNA binding protein. The biological significance of these characteristics is discussed.
细菌表达的爱泼斯坦-巴尔病毒（EBV）脱氧核糖核酸酶（DNase）被纯化至98%的纯度，并用作表征该酶活性的来源。EBV DNase对DNA的完全消化产生5'-单磷酸核苷作为最终产物。在反应的对数期，EBV DNase对双链DNA（dsDNA）进行持续性作用，但对单链DNA（ssDNA）进行分布式作用。5'至3'和3'至5'的核酸外切酶活性均存在，尽管前者的活性强10倍。当使用具有5'-突出端、平端或3'-突出端的底物时，DNase释放末端标记核苷酸的情况未观察到显著差异。以超螺旋质粒DNA为底物时，EBV DNase也被证明具有核酸内切酶活性。在pBS-TR（一种含有一份EBV末端重复序列的pBlueScript载体）上定位了两个优先的dsDNA切割位点；两者均位于载体序列中。最后，一种N端截短的EBV主要DNA结合蛋白，而不是EA-D，被证明可抑制EBV DNase活性。这种抑制作用可能是由于EBV DNase与主要DNA结合蛋白之间直接的蛋白质-蛋白质相互作用。讨论了这些特性的生物学意义。
