title: Screening natural products for inhibitors of quinone reductase-2 using ultrafiltration LC-MS.
abstract: Inhibitors of quinone reductase-2 (NQO2; QR-2) can have antimalarial activity and antitumor activities or can function as chemoprevention agents by preventing the metabolic activation of toxic quinones such as menadione. To expedite the search for new natural product inhibitors of QR-2, we developed a screening assay based on ultrafiltration liquid chromatography-mass spectrometry that is compatible with complex samples such as bacterial or botanical extracts. Human QR-2 was prepared recombinantly, and the known QR-2 inhibitor, resveratrol, was used as a positive control and as a competitive ligand to eliminate false positives. Ultrafiltration LC-MS screening of extracts of marine sediment bacteria resulted in the discovery of tetrangulol methyl ether as an inhibitor of QR-2. When applied to the screening of hop extracts from the botanical, Humulus lupulus L., xanthohumol and xanthohumol D were identified as ligands of QR-2. Inhibition of QR-2 by these ligands was confirmed using a functional enzyme assay. Furthermore, binding of xanthohumol and xanthohumol D to the active site of QR-2 was confirmed using X-ray crystallography. Ultrafiltration LC-MS was shown to be a useful assay for the discovery of inhibitors of QR-2 in complex matrixes such as extracts of bacteria and botanicals.
authors.affiliation: Department of Medicinal Chemistry and Pharmacognosy, University of Illinois College of Pharmacy, Chicago, Illinois 60612, USA.
authors.forename: Yongsoo
authors.initials: Y
authors.lastname: Choi
authors.forename: Katherine
authors.initials: K
authors.lastname: Jermihov
authors.forename: Sang-Jip
authors.initials: SJ
authors.lastname: Nam
authors.forename: Megan
authors.initials: M
authors.lastname: Sturdy
authors.forename: Katherine
authors.initials: K
authors.lastname: Maloney
authors.forename: Xi
authors.initials: X
authors.lastname: Qiu
authors.forename: Lucas R
authors.initials: LR
authors.lastname: Chadwick
authors.forename: Matthew
authors.initials: M
authors.lastname: Main
authors.forename: Shao-Nong
authors.initials: SN
authors.lastname: Chen
authors.forename: Andrew D
authors.initials: AD
authors.lastname: Mesecar
authors.forename: Norman R
authors.initials: NR
authors.lastname: Farnsworth
authors.forename: Guido F
authors.initials: GF
authors.lastname: Pauli
authors.forename: William
authors.initials: W
authors.lastname: Fenical
authors.forename: John M
authors.initials: JM
authors.lastname: Pezzuto
authors.forename: Richard B
authors.initials: RB
authors.lastname: van Breemen
authors.forename: Richard R
authors.initials: RR
authors.lastname: van Breemen
mesh_terms: D001688:Biological Products / Q000032:analysis* / Q000494:pharmacology; D002853:Chromatography, Liquid / Q000379:methods*; D018360:Crystallography, X-Ray; D004353:Drug Evaluation, Preclinical; D004791:Enzyme Inhibitors / Q000032:analysis* / Q000494:pharmacology; D005374:Filtration; D005419:Flavonoids / Q000032:analysis* / Q000494:pharmacology; D006801:Humans; D027582:Humulus / Q000737:chemistry*; D013058:Mass Spectrometry / Q000379:methods*; D008958:Models, Molecular; D015394:Molecular Structure; D016660:NAD(P)H Dehydrogenase (Quinone) / Q000037:antagonists & inhibitors*; D011427:Propiophenones / Q000032:analysis / Q000494:pharmacology
chemical_list: D001688:Biological Products; D004791:Enzyme Inhibitors; D005419:Flavonoids; D011427:Propiophenones; C558154:xanthohumol D; D016660:NAD(P)H Dehydrogenase (Quinone); C104536:xanthohumol