Superfamily-wide portrait of serine hydrolase inhibition achieved by library-versus-library screening
Superfamily-wide portrait of serine hydrolase inhibition achieved by library-versus-library screening
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DOI：
10.1073/pnas.1011663107
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发表时间：
2010-12-07
影响因子：
11.1
通讯作者：
Cravatt, Benjamin F.
中科院分区：
文献类型：
--
作者：
Bachovchin, Daniel A.;Ji, Tianyang;Cravatt, Benjamin F.
丝氨酸水解酶是哺乳动物体内最大、种类最多的一类酶。它们在几乎所有的生理过程中发挥着重要作用，并被药物靶向治疗糖尿病、肥胖和神经退行性疾病等疾病。尽管如此，我们缺乏对大多数110+预测的哺乳动物代谢SH的生物学理解，这在很大程度上是因为缺乏评估其生化活性的测定方法，并且缺乏选择性抑制剂来探测其在生命系统中的功能。我们在这里表明，绝大多数（>80%）的哺乳动物代谢SH可以被标记在蛋白质组中的一个单一的，活性位点定向的氟膦酸酯探针。我们利用这种通用的基于活性的测定在库对库的格式筛选70+ SH对140+结构不同的氨基甲酸酯。铅抑制剂被发现的相似的40%的筛选的酶，包括许多不良特征的SH。全球概况确定的氨基甲酸酯抑制剂之间的歧视高度序列相关的SH，相反，酶，尽管缺乏序列同源性，共享抑制剂敏感性概况。这些发现表明，序列相关性不是SH超家族内共享药理学的强预测因子。最后，我们表明，铅氨基甲酸酯抑制剂可以被优化成药理学探针，在体内以高特异性抑制单个SH。
Serine hydrolases (SHs) are one of the largest and most diverse enzyme classes in mammals. They play fundamental roles in virtually all physiological processes and are targeted by drugs to treat diseases such as diabetes, obesity, and neurodegenerative disorders. Despite this, we lack biological understanding for most of the 110+ predicted mammalian metabolic SHs, in large part because of a dearth of assays to assess their biochemical activities and a lack of selective inhibitors to probe their function in living systems. We show here that the vast majority (>80%) of mammalian metabolic SHs can be labeled in proteomes by a single, active site-directed fluorophosphonate probe. We exploit this universal activity-based assay in a library-versus-library format to screen 70+ SHs against 140+ structurally diverse carbamates. Lead inhibitors were discovered for similar to 40% of the screened enzymes, including many poorly characterized SHs. Global profiles identified carbamate inhibitors that discriminate among highly sequence-related SHs and, conversely, enzymes that share inhibitor sensitivity profiles despite lacking sequence homology. These findings indicate that sequence relatedness is not a strong predictor of shared pharmacology within the SH superfamily. Finally, we show that lead carbamate inhibitors can be optimized into pharmacological probes that inactivate individual SHs with high specificity in vivo.