title: Bioluminescent method for assaying multiple semicarbazide-sensitive amine oxidase (SSAO) family members in both 96- and 384-well formats.
abstract: Vascular adhesion protein-1 (VAP-1), also known as semicarbazide-sensitive amine oxidase (SSAO) or copper-containing amine oxidase (AOC3, EC 1.4.3.6), catalyzes oxidative deamination of primary amines. One endogenous substrate has recently been described (Siglec 10), and although its mechanism of action in vivo is not completely understood, it is suggested to play a role in immune cell trafficking, making it a target of interest for autoimmune and inflammatory diseases. Much of the enzymology performed around this target has been conducted with absorbance, fluorescent, or radiometric formats that can have some limitations for high-throughput screening and subsequent compound profiling. The authors present the use of a bioluminescent assay, originally developed for monoamine oxidase enzymes, in a high-throughput format. It can be used for related SSAOs such as AOC1 given their substrate similarity with VAP-1. The authors also demonstrate that it is compatible with different sources of VAP-1, both purified recombinant and VAP-1 overexpressed on live cells.
authors.affiliation: Boehringer Ingelheim Pharmaceuticals, Ridgefield, CT 06877, USA.
authors.forename: Gregory W
authors.initials: GW
authors.lastname: Peet
authors.forename: Susan
authors.initials: S
authors.lastname: Lukas
authors.forename: Melissa
authors.initials: M
authors.lastname: Hill-Drzewi
authors.forename: Leslie
authors.initials: L
authors.lastname: Martin
authors.forename: Irina V
authors.initials: IV
authors.lastname: Rybina
authors.forename: Teresa
authors.initials: T
authors.lastname: Roma
authors.forename: Alycia
authors.initials: A
authors.lastname: Shoultz
authors.forename: Xiang
authors.initials: X
authors.lastname: Zhu
authors.forename: Daniela
authors.initials: D
authors.lastname: Cazacu
authors.forename: Anthony
authors.initials: A
authors.lastname: Kronkaitis
authors.forename: Alistair
authors.initials: A
authors.lastname: Baptiste
authors.forename: Brian C
authors.initials: BC
authors.lastname: Raudenbush
authors.forename: E Michael
authors.initials: EM
authors.lastname: August
authors.forename: Louise Kelly
authors.initials: LK
authors.lastname: Modis
mesh_terms: D006631:Amine Oxidase (Copper-Containing) / Q000037:antagonists & inhibitors / Q000378:metabolism*; D000818:Animals; D016466:CHO Cells; D002461:Cell Line, Transformed; D006224:Cricetinae; D004789:Enzyme Activation / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology; D057809:HEK293 Cells; D057166:High-Throughput Screening Assays; D006801:Humans; D020128:Inhibitory Concentration 50; D008163:Luminescent Measurements; D051379:Mice; D051381:Rats
chemical_list: D004791:Enzyme Inhibitors; D006631:Amine Oxidase (Copper-Containing)