Mesopone cytochrome c peroxidase: Functional model of heme oxygenated oxidases

Chad E. Immoos, B. Bhaskar, Michael S. Cohen, Tiffany P. Barrows, Patrick J. Farmer, T. L. Poulos

Research output: Contribution to journalArticle

5 Scopus citations

Abstract

The effect of heme ring oxygenation on enzyme structure and function has been examined in a reconstituted cytochrome c peroxidase. Oxochlorin derivatives were formed by OsO4 treatment of mesoporphyrin followed by acid-catalyzed pinacol rearrangement. The northern oxochlorin isomers were isolated by chromatography, and the regio-isomers assignments determined by 2D COSY and NOE 1H NMR. The major isomer, 4-mesoporphyrinone (Mp), was metallated with FeCl2 and reconstituted into cytochrome c peroxidase (CcP) forming a hybrid green protein, MpCcP. The heme-altered enzyme has 99% wild-type peroxidase activity with cytochrome c. EPR spectroscopy of MpCcP intermediate compound I verifies the formation of the Trp191 radical similar to wild-type CcP in the reaction cycle. Peroxidase activity with small molecules is varied: guaiacol turnover increases approximately five-fold while that with ferrocyanide is ∼85% of native. The electron-withdrawing oxo-substitutents on the cofactor cause a ∼60-mV increase in FeIII/FeII reduction potential. The present investigation represents the first structural characterization of an oxochlorin protein with X-ray intensity data collected to 1.70 Å. Although a mixture of R- and S-mesopone isomers of the FeMP cofactor was used during heme incorporation into the apo-protein, only the S-isomer is found in the crystallized protein.

Original languageEnglish (US)
Pages (from-to)635-643
Number of pages9
JournalJournal of Inorganic Biochemistry
Volume91
Issue number4
DOIs
StatePublished - Sep 20 2002

Keywords

  • Cytochrome c peroxidase
  • Heme cd model
  • Reconstituted oxochlorin

ASJC Scopus subject areas

  • Biochemistry
  • Inorganic Chemistry

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