Klitgaard Rasmus N, Løbner-Olesen Anders
Department of Biology, Faculty of Science, University of Copenhagen, Copenhagen, Denmark.
Curr Drug Discov Technol. 2019;16(3):272-277. doi: 10.2174/1570163815666180423115514.
One of many strategies to overcome antibiotic resistance is the discovery of compounds targeting cellular processes, which have not yet been exploited.
Using various genetic tools, we constructed a novel high throughput, cellbased, fluorescence screen for inhibitors of chromosome replication initiation in bacteria.
The screen was validated by expression of an intra-cellular cyclic peptide interfering with the initiator protein DnaA and by over-expression of the negative initiation regulator SeqA. We also demonstrated that neither tetracycline nor ciprofloxacin triggers a false positive result. Finally, 400 extracts isolated mainly from filamentous actinomycetes were subjected to the screen.
We concluded that the presented screen is applicable for identifying putative inhibitors of DNA replication initiation in a high throughput setup.
克服抗生素耐药性的众多策略之一是发现针对尚未被利用的细胞过程的化合物。
我们使用各种遗传工具构建了一种新型的高通量、基于细胞的荧光筛选方法，用于筛选细菌染色体复制起始的抑制剂。
通过表达干扰起始蛋白DnaA的细胞内环肽以及过表达负向起始调节因子SeqA对该筛选方法进行了验证。我们还证明四环素和环丙沙星均未引发假阳性结果。最后，对主要从丝状放线菌中分离出的400种提取物进行了筛选。
我们得出结论，所展示的筛选方法适用于在高通量设置中鉴定DNA复制起始的假定抑制剂。
