title: The mechanism of formation of N-formylkynurenine by heme dioxygenases.
abstract: Heme dioxygenases catalyze the oxidation of L-tryptophan to N-formylkynurenine (NFK), the first and rate-limiting step in tryptophan catabolism. Although recent progress has been made on early stages in the mechanism, there is currently no experimental data on the mechanism of product (NFK) formation. In this work, we have used mass spectrometry to examine product formation in a number of dioxygenases. In addition to NFK formation (m/z = 237), the data identify a species (m/z = 221) that is consistent with insertion of a single atom of oxygen into the substrate during O(2)-driven turnover. The fragmentation pattern for this m/z = 221 species is consistent with a cyclic amino acetal structure; independent chemical synthesis of the 3a-hydroxypyrroloindole-2-carboxylic acid compound is in agreement with this assignment. Labeling experiments with (18)O(2) confirm the origin of the oxygen atom as arising from O(2)-dependent turnover. These data suggest that the dioxygenases use a ring-opening mechanism during NFK formation, rather than Criegee or dioxetane mechanisms as previously proposed.
authors: Department of Biochemistry, University of Leicester, Lancaster Road, Leicester LE1 9HN, United Kingdom.
authors: Jaswir
authors: J
authors: Basran
authors: Igor
authors: I
authors: Efimov
authors: Nishma
authors: N
authors: Chauhan
authors: Sarah J
authors: SJ
authors: Thackray
authors: James L
authors: JL
authors: Krupa
authors: Graham
authors: G
authors: Eaton
authors: Gerry A
authors: GA
authors: Griffith
authors: Christopher G
authors: CG
authors: Mowat
authors: Sandeep
authors: S
authors: Handa
authors: Emma Lloyd
authors: EL
authors: Raven
mesh_terms: D006418:Heme / Q000378:metabolism; D006801:Humans; D050503:Indoleamine-Pyrrole 2,3,-Dioxygenase / Q000378:metabolism*; D007737:Kynurenine / Q000031:analogs & derivatives* / Q000378:metabolism; D013058:Mass Spectrometry; D010100:Oxygen / Q000378:metabolism; D014366:Tryptophan Oxygenase / Q000378:metabolism*; D016959:Xanthomonas campestris / Q000201:enzymology
chemical_list: D050503:Indoleamine-Pyrrole 2,3,-Dioxygenase; C007772:N'-formylkynurenine; D007737:Kynurenine; D006418:Heme; D014366:Tryptophan Oxygenase; D010100:Oxygen