title: Bacterial expression of human kynurenine 3-monooxygenase: solubility, activity, purification.
abstract: Kynurenine 3-monooxygenase (KMO) is an enzyme central to the kynurenine pathway of tryptophan metabolism. KMO has been implicated as a therapeutic target in several disease states, including Huntington's disease. Recombinant human KMO protein production is challenging due to the presence of transmembrane domains, which localise KMO to the outer mitochondrial membrane and render KMO insoluble in many in vitro expression systems. Efficient bacterial expression of human KMO would accelerate drug development of KMO inhibitors but until now this has not been achieved. Here we report the first successful bacterial (Escherichia coli) expression of active FLAG™-tagged human KMO enzyme expressed in the soluble fraction and progress towards its purification.
authors: Drug Discovery Core, University/BHF Centre for Cardiovascular Science, Queen's Medical Research Institute, The University of Edinburgh, 47 Little France Crescent, Edinburgh EH16 4TJ, United Kingdom. Electronic address: Kris.Wilson@ed.ac.uk.
authors: K
authors: K
authors: Wilson
authors: MRC Centre for Inflammation Research, Queen's Medical Research Institute, The University of Edinburgh, 47 Little France Crescent, Edinburgh EH16 4TJ, United Kingdom. Electronic address: Damian.Mole@ed.ac.uk.
authors: D J
authors: DJ
authors: Mole
authors: Drug Discovery Core, University/BHF Centre for Cardiovascular Science, Queen's Medical Research Institute, The University of Edinburgh, 47 Little France Crescent, Edinburgh EH16 4TJ, United Kingdom. Electronic address: m.binnie@ed.ac.uk.
authors: M
authors: M
authors: Binnie
authors: Mass Spectrometry Core, University/BHF Centre for Cardiovascular Science, Queen's Medical Research Institute, The University of Edinburgh, 47 Little France Crescent, Edinburgh EH16 4TJ, United Kingdom. Electronic address: nhomer@staffmail.ed.ac.uk.
authors: N Z M
authors: NZ
authors: Homer
authors: MRC Centre for Inflammation Research, Queen's Medical Research Institute, The University of Edinburgh, 47 Little France Crescent, Edinburgh EH16 4TJ, United Kingdom. Electronic address: xzheng@staffmail.ed.ac.uk.
authors: X
authors: X
authors: Zheng
authors: Drug Discovery Core, University/BHF Centre for Cardiovascular Science, Queen's Medical Research Institute, The University of Edinburgh, 47 Little France Crescent, Edinburgh EH16 4TJ, United Kingdom. Electronic address: byard1@staffmail.ed.ac.uk.
authors: B A
authors: BA
authors: Yard
authors: MRC Centre for Inflammation Research, Queen's Medical Research Institute, The University of Edinburgh, 47 Little France Crescent, Edinburgh EH16 4TJ, United Kingdom. Electronic address: John.Iredale@ed.ac.uk.
authors: J P
authors: JP
authors: Iredale
authors: School of Biological Sciences and School of Biomedical Sciences, University of Edinburgh, CH Waddington Building, The University of Edinburgh Kings Buildings, Mayfield Road, Edinburgh EH9 3JD, United Kingdom. Electronic address: Manfred.Auer@ed.ac.uk.
authors: M
authors: M
authors: Auer
authors: Drug Discovery Core, University/BHF Centre for Cardiovascular Science, Queen's Medical Research Institute, The University of Edinburgh, 47 Little France Crescent, Edinburgh EH16 4TJ, United Kingdom. Electronic address: scott.webster@ed.ac.uk.
authors: S P
authors: SP
authors: Webster
mesh_terms: D004926:Escherichia coli / Q000235:genetics; D006639:Histidine; D006801:Humans; D007700:Kinetics; D050601:Kynurenine 3-Monooxygenase / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism*; D053858:Metabolic Networks and Pathways; D009842:Oligopeptides; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism*; D012995:Solubility
chemical_list: D009842:Oligopeptides; D011993:Recombinant Fusion Proteins; D006639:Histidine; C085706:FLAG peptide; D050601:Kynurenine 3-Monooxygenase
keywords: 3-Hydroxykynurenine; Kynurenine 3-monooxygenase; Purification; Solubility