biostudies-literatureMcHenry MWU.S. Department of Health & Human Services | NIH | National Cancer Institute (NCI)NIAID NIH HHSU.S. Department of Health & Human Services | NIH | National Cancer InstituteNHLBI NIH HHSU.S. Department of Health & Human Services | NIH | National Heart, Lung, and Blood InstituteNCI NIH HHSU.S. Department of Health & Human Services | NIH | National Heart, Lung, and Blood Institute (NHLBI)NIGMS NIH HHSNational Science Foundation1022-1032https://www.ebi.ac.uk/biostudies/studies/S-EPMC11252247biostudies-literatureUnknown20(8)BCL-2-associated X protein (BAX) is a promising therapeutic target for activating or restraining apoptosis in diseases of pathologic cell survival or cell death, respectively. In response to cellular stress, BAX transforms from a quiescent cytosolic monomer into a toxic oligomer that permeabilizes the mitochondria, releasing key apoptogenic factors. The mitochondrial lipid trans-2-hexadecenal (t-2-hex) sensitizes BAX activation by covalent derivatization of cysteine 126 (C126). In this study, we performed a disulfide tethering screen to discover C126-reactive molecules that modulate BAX activity. We identified covalent BAX inhibitor 1 (CBI1) as a compound that selectively derivatizes BAX at C126 and inhibits BAX activation by triggering ligands or point mutagenesis. Biochemical and structural analyses revealed that CBI1 can inhibit BAX by a dual mechanism of action: conformational constraint and competitive blockade of lipidation. These data inform a pharmacologic strategy for suppressing apoptosis in diseases of unwanted cell death by covalent targeting of BAX C126.Nature chemical biologyCovalent inhibition of pro-apoptotic BAX.PMC11252247T32 HL007574R01 AI0702925T32HL007574R35 CA197583F30 CA275293R35CA197583R01 GM067945Engen JRMcHenry MWYang KShi PWalensky LDGygi MACamara CMWales TECohen DTGygi SPRettenmaier TJAdhikary UWells JAfalseCovalent inhibition of pro-apoptotic BAX.BCL-2-associated X protein (BAX) is a promising therapeutic target for activating or restraining apoptosis in diseases of pathologic cell survival or cell death, respectively. In response to cellular stress, BAX transforms from a quiescent cytosolic monomer into a toxic oligomer that permeabilizes the mitochondria, releasing key apoptogenic factors. The mitochondrial lipid trans-2-hexadecenal (t-2-hex) sensitizes BAX activation by covalent derivatization of cysteine 126 (C126). In this study, we performed a disulfide tethering screen to discover C126-reactive molecules that modulate BAX activity. We identified covalent BAX inhibitor 1 (CBI1) as a compound that selectively derivatizes BAX at C126 and inhibits BAX activation by triggering ligands or point mutagenesis. Biochemical and structural analyses revealed that CBI1 can inhibit BAX by a dual mechanism of action: conformational constraint and competitive blockade of lipidation. These data inform a pharmacologic strategy for suppressing apoptosis in diseases of unwanted cell death by covalent targeting of BAX C126.2024-01-01T00:00:00Z2024 Aug2026-03-18T13:35:04.65Z2025-08-18T09:54:19.607ZS-EPMC112522473823358410.1038/s41589-023-01537-6