title: Tryptophan residues are targets in hypothiocyanous acid-mediated protein oxidation.
abstract: Myeloperoxidase, released by activated phagocytes, forms reactive oxidants by catalysing the reaction of halide and pseudo-halide ions with H(2)O(2). These oxidants have been linked to tissue damage in a range of inflammatory diseases. With physiological levels of halide and pseudo-halide ions, similar amounts of HOCl (hypochlorous acid) and HOSCN (hypothiocyanous acid) are produced by myeloperoxidase. Although the importance of HOSCN in initiating cellular damage via thiol oxidation is becoming increasingly recognized, there are limited data on the reactions of HOSCN with other targets. In the present study, the products of the reaction of HOSCN with proteins has been studied. With albumin, thiols are oxidized preferentially forming unstable sulfenyl thiocyanate derivatives, as evidenced by the reversible incorporation of (14)C from HOS(14)CN. On consumption of the HSA (human serum albumin) free thiol group, the formation of stable (14)C-containing products and oxidation of tryptophan residues are observed. Oxidation of tryptophan residues is observed on reaction of HOSCN with other proteins (including myoglobin, lysozyme and trypsin inhibitor), but not free tryptophan, or tryptophan-containing peptides. Peptide mass mapping studies with HOSCN-treated myoglobin, showed the addition of two oxygen atoms on either Trp(7) or Trp(14) with equimolar or less oxidant, and the addition of a further two oxygen atoms to the other tryptophan with higher oxidant concentrations (> or = 2-fold). Tryptophan oxidation was observed on treating myoglobin with HOSCN in the presence of glutathione and ascorbate. Thus tryptophan residues are likely to be favourable targets for the reaction in biological systems, and the oxidation products formed may be useful biomarkers of HOSCN-mediated protein oxidation.
authors.affiliation: The Heart Research Institute, 114 Pyrmont Bridge Road, Camperdown, Sydney, NSW 2050, Australia. hawkinsc@hri.org.au
authors.forename: Clare L
authors.initials: CL
authors.lastname: Hawkins
authors.forename: David I
authors.initials: DI
authors.lastname: Pattison
authors.forename: Naomi R
authors.initials: NR
authors.lastname: Stanley
authors.forename: Michael J
authors.initials: MJ
authors.lastname: Davies
mesh_terms: D000595:Amino Acid Sequence; D000975:Antioxidants / Q000378:metabolism; D001205:Ascorbic Acid / Q000378:metabolism; D005456:Fluorescent Dyes / Q000378:metabolism; D005978:Glutathione / Q000378:metabolism; D006801:Humans; D006997:Hypochlorous Acid / Q000737:chemistry / Q000378:metabolism; D008969:Molecular Sequence Data; D015394:Molecular Structure; D009211:Myoglobin / Q000378:metabolism; D010084:Oxidation-Reduction; D010446:Peptide Fragments / Q000737:chemistry / Q000378:metabolism; D010449:Peptide Mapping; D011489:Protein Denaturation; D012709:Serum Albumin / Q000737:chemistry / Q000378:metabolism; D013438:Sulfhydryl Compounds / Q000737:chemistry; D013861:Thiocyanates / Q000737:chemistry / Q000378:metabolism*; D014364:Tryptophan / Q000737:chemistry / Q000378:metabolism*
chemical_list: D000975:Antioxidants; D005456:Fluorescent Dyes; D009211:Myoglobin; D010446:Peptide Fragments; D012709:Serum Albumin; D013438:Sulfhydryl Compounds; D013861:Thiocyanates; C517667:hypothiocyanous acid; D006997:Hypochlorous Acid; D014364:Tryptophan; D005978:Glutathione; D001205:Ascorbic Acid