Aspergillus flavus squalene synthase as an antifungal target: Expression, activity, and inhibition
Aspergillus flavus squalene synthase as an antifungal target: Expression, activity, and inhibition
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黄曲霉角鲨烯合酶作为抗真菌靶标：表达、活性和抑制
DOI：
10.1016/j.bbrc.2019.03.070
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发表时间：
2019
影响因子：
3.1
通讯作者：
Guo Rey-Ting
中科院分区：
文献类型：
--
作者：
Song Junfeng;Shang Na;Baig Noman;Yao Jiaqi;Shin Christopher;Kim Boo Kyun;Li Qian;Malwal Satish R.;Oldfield Eric;Feng Xinxin;Guo Rey-Ting
侵袭性曲霉病（IA）是一种危及生命的疾病，影响免疫功能低下的个人。IA的标准治疗，包括多烯类和唑类，具有高毒性和新出现的耐药性，因此需要开发具有新作用机制的新抗真菌剂。麦角甾醇生物合成是抗真菌药物的经典靶标，角鲨烯合酶（SQS）催化麦角甾醇生物合成的第一个关键步骤。使SQS在抗真菌发展的背景下成为感兴趣的。在这里，我们克隆，表达，纯化和表征SQS从病原菌黄曲霉（AfSQS），证实它产生角鲨烯。为了确定针对AfSQS的潜在线索，我们测试了已知的角鲨烯合成酶抑制剂，萨拉戈萨酸和膦磺酸盐BPH-652，发现它们是有效的抑制剂。然后，我们筛选了来自国家癌症研究所（NCI）多样性集V的744种化合物的库，用于抑制活性。鉴定了20个命中，并使用剂量-反应曲线测定了IC 50值。排除干扰测定的14种化合物，并分析剩余6种化合物的药物相似性，得到一种化合物雷公藤红素，其AfSQS IC 50值为830 nM。酶抑制动力学显示雷公藤红素以非竞争性方式与AfSQS结合，但不共价结合。由于雷公藤红素还已知通过抑制黄素依赖性单加氧酶铁载体A（SidA，在铁饥饿条件下）来抑制高度致病的烟曲霉的生长，因此它可能是用于抗真菌开发的有希望的多靶点先导物。
Invasive aspergillosis (IA) is a life-threatening disease impacting immunocompromised individuals. Standard treatments of IA, including polyenes and azoles, suffer from high toxicity and emerging resistance, leading to the need to develop new antifungal agents with novel mechanisms of action. Ergosterol biosynthesis is a classic target for antifungals, and squalene synthase (SQS) catalyzes the first committed step in ergosterol biosynthesis inAspergillusspp. making SQS of interest in the context of antifungal development. Here, we cloned, expressed, purified and characterized SQS from the pathogenAspergillus flavus(AfSQS), confirming that it produced squalene. To identify potential leads targeting AfSQS, we tested known squalene synthase inhibitors, zaragozic acid and the phosphonosulfonate BPH-652, finding that they were potent inhibitors. We then screened a library of 744 compounds from the National Cancer Institute (NCI) Diversity Set V for inhibition activity. 20 hits were identified and IC50values were determined using dose-response curves. 14 compounds that interfered with the assay were excluded and the remaining 6 compounds were analyzed for drug-likeness, resulting in one compound, celastrol, which had an AfSQS IC50value of 830 nM. Enzyme inhibition kinetics revealed that celastrol binds to AfSQS in a noncompetitive manner, but did not bind covalently. Since celastrol is also known to inhibit growth of the highly virulentAspergillus fumigatusby inhibiting flavin-dependent monooxygenase siderophore A (SidA, under iron starvation conditions), it may be a promising multi-target lead for antifungal development.