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Ketamine is a non-competitive channel blocker of N-methyl-d-aspartate (NMDA) receptors.sup.1. A single sub-anaesthetic dose of ketamine produces rapid (within hours) and long-lasting antidepressant effects in patients who are resistant to other antidepressants.sup.2,3. Ketamine is a racemic mixture of S- and R-ketamine enantiomers, with S-ketamine isomer being the more active antidepressant.sup.4. Here we describe the cryo-electron microscope structures of human GluN1-GluN2A and GluN1-GluN2B NMDA receptors in complex with S-ketamine, glycine and glutamate. Both electron density maps uncovered the binding pocket for S-ketamine in the central vestibule between the channel gate and selectivity filter. Molecular dynamics simulation showed that S-ketamine moves between two distinct locations within the binding pocket. Two amino acids--leucine 642 on GluN2A (homologous to leucine 643 on GluN2B) and asparagine 616 on GluN1--were identified as key residues that form hydrophobic and hydrogen-bond interactions with ketamine, and mutations at these residues reduced the potency of ketamine in blocking NMDA receptor channel activity. These findings show structurally how ketamine binds to and acts on human NMDA receptors, and pave the way for the future development of ketamine-based antidepressants. Structures of ketamine bound to human NMDA receptors show how ketamine inhibits receptor activity., Author(s): Youyi Zhang [sup.1] [sup.2] , Fei Ye [sup.3] , Tongtong Zhang [sup.1] [sup.2] , Shiyun Lv [sup.1] [sup.2] , Liping Zhou [sup.2] [sup.4] , Daohai Du [sup.4] , He [...]

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 Z factor of 0.71. The natural product (-)-epigallocatechin gallate (EGCG) was found to be a noncompetitive inhibitor of NADK with K i = 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 K d 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., Competing Interests: The authors declare no competing financial interest., (© 2022 American Chemical Society.)