Identification of Inhibitors of the Antibiotic-Resistance Target New Delhi Metallo-β-lactamase 1 by both Nanoelectrospray Ionization Mass Spectrometry and Ultrafiltration Liquid Chromatography/Mass Spectrometry Approaches
Identification of Inhibitors of the Antibiotic-Resistance Target New Delhi Metallo-β-lactamase 1 by both Nanoelectrospray Ionization Mass Spectrometry and Ultrafiltration Liquid Chromatography/Mass Spectrometry Approaches
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DOI：
10.1021/ac401732d
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发表时间：
2013-08-20
影响因子：
7.4
通讯作者：
Shui, Wenqing
中科院分区：
文献类型：
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作者：
Chen, Xin;Li, Lixin;Shui, Wenqing
基于质谱的平台在发现与作为候选药物的治疗靶点结合的配体方面取得了越来越多的成功。我们建立了纳米电喷雾电离质谱（nanoESI-MS）测定和超滤液相色谱/质谱（LC/MS）测定，以鉴定导致全球抗生素耐药性的新德里金属β-内酰胺酶1（NDM-1）的新配体。为了减轻疏水化合物的非特异性结合并消除基于间接LC/MS的测定中通常遇到的假阳性，我们在对照中引入了阻断蛋白，这显著提高了间接方法的选择性和准确性。首次对两种基于MS的方法进行并排比较，进一步揭示了间接方法的独特优势，包括更好的重现性和干扰耐受性。此外，从小分子片段的混合物中钓出有效配体的成功证明了基于间接LC/MS的方法用于构建针对组合文库或天然产物提取物的稳健筛选平台的巨大潜力。更重要的是，通过结合基于MS的分析、酶活性测定、竞争实验和结构模拟的结果，我们发现了一种新的化合物作为靶向NDM-1的有希望的候选药物。
Mass spectrometry-based platforms have gained increasing success in discovery of ligands bound to therapeutic targets as drug candidates. We established both a nanoelectrospray ionization mass spectrometry (nanoESI-MS) assay and an ultrafiltration liquid chromatography/mass spectrometry (LC/MS) assay to identify new ligands for New Delhi metallo-beta-lactamase 1 (NDM-1), responsible for worldwide antibiotic resistance. To alleviate nonspecific binding of hydrophobic compounds and eliminate false positives typically encountered in the indirect LC/MS-based assay, we introduced a blocking protein in the control, which remarkably enhances the selectivity and accuracy of the indirect approach. Side-by-side comparison of the two MS-based approaches for the first time further reveals unique advantages of the indirect approach, including better reproducibility and tolerance of interference. Moreover, the success of fishing out a potent ligand from a mixture of small-molecule fragments demonstrates great potential of the indirect LC/MS-based approach for constructing a robust screening platform against combinatorial libraries or natural product extracts. More importantly, by combining the results of MS-based analyses, enzymatic activity assay, competition experiments, and structural simulation, we discovered a new compound as a promising drug candidate targeting NDM-1.