Discovery of FabH/FabF inhibitors from natural products
Discovery of FabH/FabF inhibitors from natural products
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DOI：
10.1128/aac.50.2.519-526.2006
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发表时间：
2006-02-01
影响因子：
4.9
通讯作者：
Wang, J
中科院分区：
文献类型：
--
作者：
Young, K;Jayasuriya, H;Wang, J
缩合酶在11型脂肪酸的合成中是必不可少的，也是抗菌药物开发的有希望的靶标。最近，一种利用木糖诱导的质粒在金黄色葡萄球菌中表达反义RNA的新方法已被描述，但其实际机制尚未阐明。本文利用Northern印迹技术研究了反义RNA表达降低靶蛋白产量的机制。这表明，反义RNA通过靶向mRNA而在转录后发挥作用，导致5‘mRNA的降解。利用这项技术，建立了一种双板法，通过对天然产品提取物的筛选来鉴定FabF/FabH靶标细胞通透性抑制剂。对25万多种天然产物发酵液进行了筛选，并通过生化分析进行了确认，命中率为0.1%。利用这种基于全细胞机制的筛选方法，发现了所有已知的天然产物FabH和FabF抑制剂，包括蓝菌素、硫代乳霉素、硫代曲霉素和Tu3010。还发现了对苯二烯酸，这是一种新的FabF抑制剂。这些新的抑制剂在凝胶延伸试验和基于全细胞的双板试验中表现出靶向性。对马来酸C对金黄色葡萄球菌有较好的抗菌活性，约为硫代乳霉素和蓝菌素的20倍。它对包括耐甲氧西林金黄色葡萄球菌、枯草芽孢杆菌和流感嗜血杆菌在内的临床重要病原体表现出广泛的抗菌活性。
Condensing enzymes are essential in type 11 fatty acid synthesis and are promising targets for antibacterial drug discovery. Recently, a new approach using a xylose-inducible plasmid to express antisense RNA in Staphylococcus aureus has been described; however, the actual mechanism was not delineated. In this paper, the mechanism of decreased target protein production by expression of antisense RNA was investigated using Northern blotting. This revealed that the antisense RNA acts posttranscriptionally by targeting mRNA, leading to 5' mRNA degradation. Using this technology, a two-plate assay was developed in order to identify FabF/FabH target-specific cell-permeable inhibitors by screening of natural product extracts. Over 250,000 natural product fermentation broths were screened and then confirmed in biochemical assays, yielding a hit rate of 0.1%. All known natural product FabH and FabF inhibitors, including cerulenin, thiolactomycin, thiotetromycin, and Tu3010, were discovered using this whole-cell mechanism-based screening approach. Phomallenic acids, which are new inhibitors of FabF, were also discovered. These new inhibitors exhibited target selectivity in the gel elongation assay and in the whole-cell-based two-plate assay. Phomallenic acid C showed good antibacterial activity, about 20-fold better than that of thiolactomycin and cerulenin, against S. aureus. It exhibited a spectrum of antibacterial activity against clinically important pathogens including methicillin-resistant Staphylococcus aureus, Bacillus subtilis, and Haemophilus influenzae.