Propylselen inhibits cancer cell growth by targeting glutamate dehydrogenase at the NADP+ binding site

High levels of glutamate dehydrogenase (GDH) activity are associated with hypoglycemia, cancer, and Parkinson's disease. Propylselen was synthesized to investigate its mechanism of GDH inhibition in comparison with Ebselen and Epigallocatechin gallate (EGCG). Because Ebselen was found to crosslink with the peptide (AA299-341) at the active site of E.coli GDH, the Cys, Pro, and Lys residues of the corresponding peptide were mutagenized to Ala residues. Using enzyme kinetics and biomolecular interaction assays, we found that the conserved GDH P320 residue is important for propylselen binding, C321 for Ebselen binding, and K341 for EGCG binding. In addition, these 3 mutations abolished NADP(+) binding to E. coli GDH in the absence of glutamate substrate, but in the presence of glutamate, the catalytic activity of the mutants was reduced only by 2-4 fold, indicating that a substrate-induced fit mechanism exists in E. coli GDH. Furthermore, biochemical analysis showed that NADP(+) had high affinity (Kd of 77 nM) for GDH; by targeting the NADP binding site, propylselen effectively inhibited both E. coli and human GDH activity and improved anticancer activity. (C) 2018 Elsevier Inc. All rights reserved.