(-)-Epigallocatechin Gallate is a Noncompetitive Inhibitor of NAD Kinase.
Liu T, Shi W, Ding Y, Wu Q, Zhang B, Zhang N, Wang M, Du D, Zhang H, Han B, Guo D, Zheng J, Li Q, Luo C.
Nicotinamide adenine dinucleotide kinase (NADK) controls the intracellular NADPH content and provides reducing power for the synthesis of macromolecules and anti-ROS. Moreover, NADK is considered to be a synthetic lethal gene for KRAS mutations. To discover NADK-targeted probes, a high-throughput screening assay was established and optimized with a <i>Z</i> factor of 0.71. The natural product (-)-epigallocatechin gallate (EGCG) was found to be a noncompetitive inhibitor of NADK with <i>K</i> <sub>i</sub> = 3.28 ± 0.32 μΜ. The direct binding of EGCG to NADK was determined by several biophysical methods, including NMR spectroscopy, surface plasmon resonance (SPR) assay, and hydrogen-deuterium exchange mass spectrometry (HDX-MS). The SPR assay showed a <i>K</i> <sub>d</sub> of 1.78 ± 1.15 μΜ. The HDX-MS experiment showed that EGCG was bound at the non-substrate-binding sites of NADK. Besides, binding mode prediction and derivative activity analysis revealed a potential structure-activity relationship between EGCG and NADK. Furthermore, EGCG can specifically inhibit the proliferation of KRAS-mutated lung cancer cell lines without affecting KRAS wild-type lung cancer cell lines.