A predicted structure of NADPH Oxidase 1 identifies key components of ROS generation and strategies for inhibition

• Published in: PloS one Vol. 18; no. 5; p. e0285206
• Main Authors: Liu, Yezhou, Liang, Shiyu, Shi, Danfeng, Zhang, Yue, Bai, Chen, Ye, Richard D
• Format: Journal Article
• Language: English
• Published: United States Public Library of Science 03.05.2023; Public Library of Science (PLoS)
• Subjects: Algorithms; Analysis; Antibodies; Binding; Biology and Life Sciences; Biotechnology; Cell culture; Cloning; Deep learning; DNA; Domains; Drug development; Electron transfer; Electron transport; Electron transport chain; Enzymes; Epithelial cells; Epithelium; Health aspects; Heme; Inhibitors; Membranes; Microenvironments; Modelling; Molecular docking; Molecular Docking Simulation; Mutagenesis; NAD(P)H oxidase; NADH, NADPH Oxidoreductases - metabolism; NADP; NADPH Oxidase 1 - genetics; NADPH Oxidases - metabolism; Oxidase; Oxidases; Oxygen; Pathogenesis; Physical Sciences; Plasmids; Proteins; Reactive oxygen species; Reactive Oxygen Species - metabolism; Reagents; Research and Analysis Methods; Site-directed mutagenesis; Substrates; China; St Louis Missouri; United Kingdom > UK; United States > US
• ISSN: 1932-6203; 1932-6203
• DOI: 10.1371/journal.pone.0285206

NADPH oxidase 1 (NOX1) is primarily expressed in epithelial cells and responsible for local generation of reactive oxygen species (ROS). By specifically manipulating the local redox microenvironment, NOX1 actively engages in epithelial immunity, especially in colorectal and pulmonary epithelia. To unravel the structural basis of NOX1 engaged epithelial immune processes, a predicted structure model was established using RaptorX deep learning models. The predicted structure model illustrates a 6-transmembrane domain structure, a FAD binding domain, and an NADPH binding/NOXO1 interacting region. The substrate/cofactor binding scheme with respect to this proposed model highly correlates with published reports and is verified in our site-directed mutagenesis assays. An electron transport chain, from NADPH to FAD and the two heme groups, was well supported by the predicted model. Through molecular docking analysis of various small molecule NOX1 inhibitors and subsequent experimental validation, we identified pronounced active sites for potent NOX1 inhibition. Specifically, LEU60, VAL71, MET181, LEU185, HIS208, PHE211, TYR214, and TYR280 in the transmembrane domain form an active pocket for insertion of the small molecule inhibitors to inhibit electron transfer between the heme groups, thus affecting extracellular ROS generation. Altogether, our study provides structural information to help elucidate the role of NOX1 in epithelial generation of ROS and sheds light on the development of therapeutics for NOX1 related illnesses.