Peroxide-dependent formation of a covalent link between Trp51 and the heme in cytochrome c peroxidase.

TitlePeroxide-dependent formation of a covalent link between Trp51 and the heme in cytochrome c peroxidase.
Publication TypeJournal
AuthorsPipirou, Z, Guallar, V, Basran, J, Metcalfe, CL, Murphy, EJ, Bottrill, AR, Mistry, SC, Raven, EL
Year of Publication2009
Date Published2009 Apr 28
PublicationBiochemistry
Volume48
Issue16
Pagination3593-9
Publication Languageeng
ISSN Number1520-4995
KeywordsChromatography, High Pressure Liquid, Cytochrome-c Peroxidase, Heme, Molecular Structure, Oxidants, Peroxides, Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization, Tryptophan
Abstract

Ascorbate peroxidase (APX), cytochrome c peroxidase (CcP), and the catalase-peroxidases (KatG) share very similar active site structures and are distinguished from other peroxidases by the presence of a distal tryptophan residue. In KatG, this distal tryptophan forms a covalent link to an adjacent tyrosine residue, which in turn links to a methionine residue. We have previously shown [ Pipirou, Z. et al. ( 2007 ) Biochemistry 46 , 2174 - 2180 ] that reaction of APX with peroxide leads, over long time scales, to formation of a covalent link with the distal tryptophan (Trp41) in a mechanism that proceeds through initial formation of a compound I species bearing a porphyrin pi-cation radical followed by radical formation on Trp41, as implicated in the KatG enzymes. Formation of such a covalent link in CcP has never been reported, and we proposed that this could be because compound I in CcP uses Trp191 instead of a porphyrin pi-cation radical. To test this, we have examined the reactivity of the W191F variant of CcP with H(2)O(2), in which formation of a porphyrin pi-cation radical occurs. We show, using electronic spectroscopy, HPLC, and mass spectroscopy, that in W191F partial formation of a covalent link from Trp51 to the heme is observed, as in APX. Radical formation on Trp51, as seen for KatG and APX, is implicated; this is supported by QM/MM calculations. Collectively, the data show that all three members of the class I heme peroxidases can support radical formation on the distal tryptophan and that the reactivity of this radical can be controlled either by the protein structure or by the nature of the compound I intermediate.

 
 
DOI10.1021/bi802210g

http://www.ncbi.nlm.nih.gov/pubmed/19249872?dopt=Abstract

 
 
Citation Key3639
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