title: A bioluminescent assay for monoamine oxidase activity.
abstract: This article describes a novel two-step homogeneous bioluminescent assay for monoamine oxidase (MAO) that is simple, sensitive, and amenable to high-throughput screening. In the first step, MAO reacts with an aminopropylether analog of methyl ester luciferin. In the second step, a luciferin detection reagent inactivates MAO and converts the product of the first step into a luminescent signal. The amount of light produced is proportional to the amount of MAO and the time of incubation in the first step, but the luminescent signal is stable in the second step with a half-life greater than 5h. The assay has high precision, is more sensitive than current fluorescent methods, and can accurately measure the binding constants of known substrates and inhibitors. An automated screen of the Sigma-RBI Library of Pharmacologically Active Compounds (LOPAC(1280)) revealed a surprisingly high percentage of MAO inhibitors (16%) with a low false hit rate (0.9%). This implies that a significant number of compounds interact with the MAO enzymes and suggests that it is important to include MAO assays in drug metabolism studies. Other advantages of this bioluminescent assay over comparable fluorescent assays are discussed.
authors.affiliation: Promega Corporation, Madison, WI 53711, USA. mike.valley@promega.com
authors.forename: Michael P
authors.initials: MP
authors.lastname: Valley
authors.forename: Wenhui
authors.initials: W
authors.lastname: Zhou
authors.forename: Erika M
authors.initials: EM
authors.lastname: Hawkins
authors.forename: John
authors.initials: J
authors.lastname: Shultz
authors.forename: James J
authors.initials: JJ
authors.lastname: Cali
authors.forename: Tracy
authors.initials: T
authors.lastname: Worzella
authors.forename: Laurent
authors.initials: L
authors.lastname: Bernad
authors.forename: Troy
authors.initials: T
authors.lastname: Good
authors.forename: Dave
authors.initials: D
authors.lastname: Good
authors.forename: Terry L
authors.initials: TL
authors.lastname: Riss
authors.forename: Dieter H
authors.initials: DH
authors.lastname: Klaubert
authors.forename: Keith V
authors.initials: KV
authors.lastname: Wood
mesh_terms: D000818:Animals; D007700:Kinetics; D008156:Luciferases / Q000378:metabolism / Q000493:pharmacokinetics; D008163:Luminescent Measurements / Q000379:methods*; D053858:Metabolic Networks and Pathways; D008738:Methyl Ethers / Q000737:chemistry / Q000493:pharmacokinetics; D051379:Mice; D008930:Mitochondria, Liver / Q000378:metabolism; D008995:Monoamine Oxidase / Q000032:analysis* / Q000378:metabolism*; D008996:Monoamine Oxidase Inhibitors / Q000378:metabolism*; D012680:Sensitivity and Specificity
chemical_list: D008738:Methyl Ethers; D008996:Monoamine Oxidase Inhibitors; D008156:Luciferases; D008995:Monoamine Oxidase