Baggelaar Marc P, Van der Stelt Mario
Department of Bio-organic Synthesis, Leiden University, 9500, Einsteinweg 55, 2333 CC, Leiden, The Netherlands.
Methods Mol Biol. 2017;1491:161-169. doi: 10.1007/978-1-4939-6439-0_12.
Competitive activity-based protein profiling is a highly efficient chemical biology technique to determine target engagement and selectivity profiles of enzyme inhibitors in complex proteomes. Fluorophosphonate-based fluorescent inhibitors are widely used as broad-spectrum probes for serine hydrolases. However, diacylglycerol lipase-α is not labeled by fluorophosphonate-based probes. To overcome this problem, we have developed a tailor-made activity-based probe that reacts with diacylglycerol lipase-α. Here we describe a case study in which we apply competitive activity-based protein profiling using a broad-spectrum and a tailor-made activity-based probe to establish selectivity and activity profiles of inhibitors targeting diacylglycerol lipase-α in the mouse brain proteome.
基于竞争性活性的蛋白质谱分析是一种高效的化学生物学技术，用于确定复杂蛋白质组中酶抑制剂的靶点结合情况和选择性谱。基于氟膦酸酯的荧光抑制剂被广泛用作丝氨酸水解酶的广谱探针。然而，二酰基甘油脂肪酶-α不会被基于氟膦酸酯的探针标记。为了克服这个问题，我们开发了一种与二酰基甘油脂肪酶-α反应的定制活性探针。在此，我们描述了一个案例研究，其中我们使用广谱和定制的基于活性的探针进行基于竞争性活性的蛋白质谱分析，以建立针对小鼠脑蛋白质组中二酰基甘油脂肪酶-α的抑制剂的选择性和活性谱。
