title: The reactivity of heme in biological systems: autocatalytic formation of both tyrosine-heme and tryptophan-heme covalent links in a single protein architecture.
abstract: We have previously shown that introduction of an engineered Met160 residue in ascorbate peroxidase (S160M variant) leads to the formation of a covalent link between Met160 and the heme vinyl group [Metcalfe, C. L., et al. (2004) J. Am. Chem. Soc. 126, 16242-16248]. In this work, we have used electronic spectroscopy, HPLC, and mass spectrometry to show that the introduction of a tyrosine residue at the same position (S160Y variant) leads, similarly, to the formation of a heme-tyrosine covalent link in an autocatalytic reaction that also leads to formation of a second covalent link from the heme to Trp41 [Pipirou, Z., et al. (2007) Biochemistry 46, 2174-2180]. Stopped-flow and EPR data implicate the involvement of a tyrosyl radical in the reaction mechanism. The results indicate that the heme can support the formation of different types of covalent links under appropriate conditions. The generality of this idea is discussed in the context of other heme enzymes.
authors.affiliation: Department of Chemistry, Henry Wellcome Building, University of Leicester, University Road, Leicester, LE1 7RH, England.
authors.forename: Zoi
authors.initials: Z
authors.lastname: Pipirou
authors.forename: Andrew R
authors.initials: AR
authors.lastname: Bottrill
authors.forename: Dimitri A
authors.initials: DA
authors.lastname: Svistunenko
authors.forename: Igor
authors.initials: I
authors.lastname: Efimov
authors.forename: Jaswir
authors.initials: J
authors.lastname: Basran
authors.forename: Sharad C
authors.initials: SC
authors.lastname: Mistry
authors.forename: Christopher E
authors.initials: CE
authors.lastname: Cooper
authors.forename: Emma Lloyd
authors.initials: EL
authors.lastname: Raven
mesh_terms: D000595:Amino Acid Sequence; D060387:Ascorbate Peroxidases; D002384:Catalysis; D002851:Chromatography, High Pressure Liquid; D004578:Electron Spin Resonance Spectroscopy; D006418:Heme / Q000737:chemistry* / Q000378:metabolism; D006861:Hydrogen Peroxide / Q000378:metabolism; D013058:Mass Spectrometry; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010084:Oxidation-Reduction; D010544:Peroxidases / Q000737:chemistry* / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D013025:Glycine max / Q000201:enzymology; D014364:Tryptophan / Q000737:chemistry* / Q000378:metabolism; D014443:Tyrosine / Q000737:chemistry* / Q000378:metabolism
chemical_list: D011994:Recombinant Proteins; D014443:Tyrosine; D006418:Heme; D014364:Tryptophan; D006861:Hydrogen Peroxide; D010544:Peroxidases; D060387:Ascorbate Peroxidases