Affinity-Based Fluorescence Polarization Assay for High-Throughput Screening of Prolyl Hydroxylase 2 Inhibitors

Yonghua Lei; Tianhan Hu; Xingsen Wu; Yue Wu; Qichao Bao; Lianshan Zhang; Hua Xia; Haopeng Sun; Qidong You; Xiaojin Zhang

doi:10.1021/acsmedchemlett.5b00394

Affinity-Based Fluorescence Polarization Assay for High-Throughput Screening of Prolyl Hydroxylase 2 Inhibitors

ACS Med Chem Lett. 2015 Nov 9;6(12):1236-40. doi: 10.1021/acsmedchemlett.5b00394. eCollection 2015 Dec 10.

Authors

Yonghua Lei¹, Tianhan Hu¹, Xingsen Wu¹, Yue Wu¹, Qichao Bao¹, Lianshan Zhang², Hua Xia¹, Haopeng Sun¹, Qidong You¹, Xiaojin Zhang³

Affiliations

¹ Jiangsu Key Laboratory of Drug Design and Optimization, and State Key Laboratory of Natural Medicines, China Pharmaceutical University , Nanjing 210009, China.
² National Engineering and Research Center for Target Drugs , Lianyungang 222000, China.
³ Jiangsu Key Laboratory of Drug Design and Optimization, and State Key Laboratory of Natural Medicines, China Pharmaceutical University , Nanjing 210009, China ; Department of Organic Chemistry, School of Science, China Pharmaceutical University , Nanjing 210009, China.

Abstract

Prolyl hydroxylase domain 2 (PHD2) enzyme, a Fe(II) and 2-oxoglutarate (2-OG) dependent oxygenase, mediates key physiological responses to hypoxia by modulating the levels of hypoxia inducible factor 1-α (HIF1α). PHD2 has been shown to have the therapeutic potentials for conditions including anemia and ischemic disease. Currently, many activity-based assays have been developed for identifying PHD2 inhibitors. Here we report an affinity-based fluorescence polarization method using FITC-labeled HIF1α (556-574) peptide as a probe for quantitative and site-specific screening of small molecule PHD2 inhibitors.

Keywords: FITC-labeled probe; PHD2; affinity-based; fluorescence polarization assay.