Papworth Monika, Moore Michael, Isalan Mark, Minczuk Michal, Choo Yen, Klug Aaron
Medical Research Council Laboratory of Molecular Biology, Hills Road, Cambridge CB2 2QH, United Kingdom.
Proc Natl Acad Sci U S A. 2003 Feb 18;100(4):1621-6. doi: 10.1073/pnas.252773399. Epub 2003 Feb 6.
The herpes simplex virus 1 (HSV-1) replicative cycle begins by binding of the viral activator, VP16, to a set of sequences in the immediate-early (IE) gene promoters. With the aim of inhibiting this cycle, we have constructed a number of synthetic zinc-finger DNA-binding peptides by using recently reported methods. Peptides containing either three or six fingers, targeted to a viral promoter, were engineered as fusions with a KOX-1 transcription repression domain. These proteins bound to the HSV-1 IE175k (ICP4) promoter, in vitro, with nanomolar or subnanomolar binding affinity. However, in a chloramphenicol acetyltransferase reporter system, only the six-finger protein was found to repress VP16-activated transcription significantly. Thus the longer array of zinc fingers is required to compete successfully against VP16, one of the most powerful natural activators known. We found that the HSV-1 replication cycle can be partially repressed by the six-finger peptide with the viral titer reduced by 90%.
单纯疱疹病毒1型（HSV-1）的复制周期始于病毒激活剂VP16与立即早期（IE）基因启动子中的一组序列结合。为了抑制这个周期，我们使用最近报道的方法构建了一些合成锌指DNA结合肽。靶向病毒启动子的含三个或六个锌指的肽被设计成与KOX-1转录抑制结构域融合。这些蛋白质在体外以纳摩尔或亚纳摩尔的结合亲和力与HSV-1 IE175k（ICP4）启动子结合。然而，在氯霉素乙酰转移酶报告系统中，仅发现六指蛋白能显著抑制VP16激活的转录。因此，需要更长的锌指阵列才能成功对抗VP16，VP16是已知最强大的天然激活剂之一。我们发现，六指肽可部分抑制HSV-1的复制周期，病毒滴度降低90%。
