Discovery And Mechanism Of Small Molecule Inhibitors Targeting GAPDH

Backgrounds: Aerobic glycolysis,which is closely related to Warburg effect,plays an increasingly important role in metabolism,tumor and immune diseases.Glyceraldehyde3-phosphate dehydrogenase(GAPDH)is a key rate-limiting enzyme in the glycolysis pathway.It catalyzes the conversion of glyceraldehyde-3-phosphate to1,3-Bisphosphoglyceric acid,and uses NAD+ as a hydrogen receptor to produce NADH.NADH is mainly involved in material and energy metabolism in cells.It transfers energy and assists ATP production through oxidative phosphorylation in mitochondria.At present,GAPDH has been a hot topic for researchers.Significance: Current studies have shown that GAPDH is associated with many diseases,such as neurological diseases,tumors,immune diseases,metabolism diseases and so on. But,the developed GAPDH inhibitors are still has some problems in aspect of activities and drug properties.Therefore,it is extraordinary necessary to screen small molecule inhibitors of GAPDH protein.Projects: According to the principle of producing NADH in the glycolysis pathway,we screened the active compound through the absorbance colorimetry,designed the binding confirmation experiments with GAPDH for the hit compound,and then carried out cell functional studies,investigated the compound activities in animal model.Research methods: A high-throughput screening system for enzyme activity.Molecular experiments,such as in vitro protein purification,protein thermal shift,micro-thermophoresis.Cellular experiments,such as intracellular enzyme activity and animal experiments,for example,mouse survival time were carried out.Consequence: Established a new high-throughput screen assay for GAPDH and yield a natural product-PGG,it as a reversible inhibitor of GAPDH protein,PGG downregulated the GAPDH-dependent glycolysis pathway.In cell experiments,PGG could significantly inhibit the activation of macrophages and the release of pro-inflammatory factors.Also the LPS/D-Ga IN induced acute liver injury mice model indicated that PGG could prolong the survival rate.Conclusion: We found the small molecule compounds targeting GAPDH and preliminarily studied its mechanism in inflammationInnovation: A new high-throughput enzyme activity method has been established,and a new small molecular inhibitor of GAPDH with new mechanism of action has been found.