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Natural product 1,2,3,4,6-penta-O-galloyl-β-D-glucopyranose is a reversible inhibitor of glyceraldehyde 3-phosphate dehydrogenase

Wen Li1, Li-ping Liao2,3, Ning Song3,4, Yan-jun Liu2,5, Yi-luan Ding6, Yuan-yuan Zhang2, Xiao-ru Zhou2,3,7, Zhong-ya Sun2,8, Sen-hao Xiao2,3,7, Hong-bo Wang1, Jing Lu1, Nai-xia Zhang6, Hua-liang Jiang2,7, Kai-xian Chen2,7, Chuan-peng Liu8, Jie Zheng4, Ke-hao Zhao1, Cheng Luo2,3,7,8,9
1 School of Pharmacy, Key Laboratory of Molecular Pharmacology and Drug Evaluation (Yantai University), Ministry of Education, Collaborative Innovation Center of Advanced Drug Delivery System and Biotech Drugs in Universities of Shandong, Yantai University, Yantai 264005, China
2 Drug Discovery and Design Center, the Center for Chemical Biology, State Key Laboratory of Drug Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai 201203, China
3 University of Chinese Academy of Sciences, Beijing 100049, China
4 Center of Immunological Diseases, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai 201203, China
5 Department of Gastroenterology, Xinhua Hospital, School of Medicine, Shanghai Jiaotong University, Shanghai 201210, China
6 Department of Analytical Chemistry, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai 201203, China
7 School of Life Science and Technology, Shanghai Tech University, Shanghai 201210, China
8 School of Life Science and Technology, Harbin Institute of Technology, Harbin 200092, China
9 Department of Pharmacology, College of Pharmacy, Fujian Medical University, Fuzhou 350108, China
Correspondence to: Jie Zheng: jzheng@simm.ac.cn, Ke-hao Zhao: kehaozhao@gmail.com, Cheng Luo: cluo@simm.ac.cn,
DOI: 10.1038/s41401-021-00653-0
Received: 17 December 2020
Accepted: 13 March 2021
Advance online: 13 April 2021

Abstract

Aerobic glycolysis, also known as the Warburg effect, is a hallmark of cancer cell glucose metabolism and plays a crucial role in the activation of various types of immune cells. Glyceraldehyde 3-phosphate dehydrogenase (GAPDH) catalyzes the conversion of D-glyceraldehyde 3-phosphate to D-glycerate 1,3-bisphosphate in the 6th critical step in glycolysis. GAPDH exerts metabolic flux control during aerobic glycolysis and therefore is an attractive therapeutic target for cancer and autoimmune diseases. Recently, GAPDH inhibitors were reported to function through common suicide inactivation by covalent binding to the cysteine catalytic residue of GAPDH. Herein, by developing a high-throughput enzymatic screening assay, we discovered that the natural product 1,2,3,4,6-penta-O-galloyl-β-D-glucopyranose (PGG) is an inhibitor of GAPDH with Ki = 0.5 μM. PGG blocks GAPDH activity by a reversible and NAD+ and Pi competitive mechanism, suggesting that it represents a novel class of GAPDH inhibitors. In-depth hydrogen deuterium exchange mass spectrometry (HDX-MS) analysis revealed that PGG binds to a region that disrupts NAD+ and inorganic phosphate binding, resulting in a distal conformational change at the GAPDH tetramer interface. In addition, structural modeling analysis indicated that PGG probably reversibly binds to the center pocket of GAPDH. Moreover, PGG inhibits LPS- stimulated macrophage activation by specific downregulation of GAPDH-dependent glucose consumption and lactate production. In summary, PGG represents a novel class of GAPDH inhibitors that probably reversibly binds to the center pocket of GAPDH. Our study sheds new light on factors for designing a more potent and specific inhibitor of GAPDH for future therapeutic applications.
Keywords: 3 4 2 glyceraldehyde 3-phosphate dehydrogenase; 1 6-penta-O-galloyl-β-D-glucopyranose; hydrogen deuterium exchange mass spectrometry; reversible inhibitor; glycolysis

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