Institute for Research in Biomedicine, Baldiri Reixac 10, 08028 Barcelona, Spain.
Chembiochem. 2010 May 17;11(8):1115-9. doi: 10.1002/cbic.201000019.
Prolyl oligopeptidase (POP) and dipeptidyl peptidase IV (DPP IV) are serine proteases that belong to the same family of enzymes. These peptidases are relevant because of their association with the pathophysiology of serious illnesses, such as type 2 diabetes (DPP IV), and those related to cognitive disorders (POP). Several NMR-based screening methods are being used to find and validate new hit scaffolds. In particular, (19)F NMR-based screening methods have proven to be powerful tools for the discovery and development of new inhibitors. Here we present an accurate and reliable (19)F NMR-based simultaneous assay that is used to screen for new selective POP and DPP IV inhibitors in compound mixtures. This activity assay consists of the simultaneous performance of POP and DPP-IV (19)F NMR activity assays in the presence of their fluorine-containing substrates. Furthermore, the assays were conducted in the presence of 0.01 % v/v of Triton X-100, which is a detergent that disrupts micelle formation, thereby preventing unspecific aggregate-based inhibition. Finally, this (19)F NMR methodology was applied to screen for ligands in plant extracts. Our results indicate that this method allows the simultaneous and accurate identification of selective POP and DPP IV inhibitors in these compound mixtures.
脯氨酰寡肽酶（POP）和二肽基肽酶 IV（DPP IV）是属于同一酶家族的丝氨酸蛋白酶。这些肽酶与严重疾病的病理生理学有关，例如 2 型糖尿病（DPP IV）和与认知障碍有关的疾病（POP）。目前正在使用多种基于 NMR 的筛选方法来寻找和验证新的命中支架。特别是，基于 (19)F NMR 的筛选方法已被证明是发现和开发新抑制剂的有力工具。在这里，我们提出了一种准确可靠的基于 (19)F NMR 的同时测定法，用于在化合物混合物中筛选新的选择性 POP 和 DPP IV 抑制剂。该活性测定法包括在含有其含氟底物的情况下同时进行 POP 和 DPP-IV（19）F NMR 活性测定。此外，测定法在 0.01％v/v 的 Triton X-100 存在下进行，Triton X-100 是一种去污剂，可破坏胶束形成，从而防止基于非特异性聚集的抑制。最后，该 (19)F NMR 方法学用于筛选植物提取物中的配体。我们的结果表明，该方法允许在这些化合物混合物中同时且准确地鉴定选择性的 POP 和 DPP IV 抑制剂。
