Characterization of the active site topography of manganese peroxidase using mechanism based inhibitors

Conference · Mon Mar 11 04:00:00 EST 1991 · FASEB Journal (Federation of American Societies for Experimental Biology); (United States)

OSTI ID:5373625

Harris, R; Wartishi, H; Gold, M; de Montellano, P O ^[1]

---

1. Univ. of California, San Francisco (United States) Oregon Graduate Inst. of Science and Technology, Beaverton (United States)

Manganese peroxidase (MnP), extracellular heme enzyme from the lignin-degrading fungus Phanerochaete chrysosporium, normally oxidizes Mn(II) to Mn(III). MnP is rapidly and completely inactivated in a H{sub 2}O{sub 2}-dependent reaction by 2 equivalents of sodium azide. The inactivation is paralleled by formation of azidyl radicals and high yield conversion of the prosthetic heme into a mexo-azido adduct. The meso-azido enzyme is oxidized by H{sub 2}O{sub 2} to a Compound II-like species. MnP is also inactivated by phenyl-, methyl- and ethylhydrazine. The phenylhydrazine reaction is too rapid for kinetic analysis, but K{sub I} = 402 mM and k{sub inact} = 0.22 min{sup {minus}1} for the inactivation by methylhydrazine. Reaction with phenylhydrazine at pH 4.5 does not yield iron-phenyl, N-phenyl, or meso-phenyl heme adducts. Ethylhydrazine inactivates the enzyme both at pH 4.5 and 7.0 but a {delta}-meso-ethylheme product is detected only at pH 7.0. Reconstitution of apo-MnP with {delta}-meso-ethylheme yields enzyme still capable of forming a Compound II-like species yet having diminished catalytic activity. Finally, Co(II) inhibits the enzyme competitively with respect to Mn(II) but does not inhibit its inactivation by azide or the alkylhydrazines. The results argue that substrates interact with the heme edge in the vicinity of the {delta}-meso carbon. They also suggest that Mn(II) and Co(II) bind to a common site close to the {delta}-meso carbon without blocking the approach of small molecules to the heme edge. An active site model is proposed that accommodates these results.

• OSTI ID: 5373625
• Report Number(s): CONF-9104107--
• Conference Information: Journal Name: FASEB Journal (Federation of American Societies for Experimental Biology); (United States) Journal Volume: 5:4
• Country of Publication: United States
• Language: English

Related Subjects

560300* -- Chemicals Metabolism & Toxicology
63 RADIATION, THERMAL, AND OTHER ENVIRON. POLLUTANT EFFECTS ON LIVING ORGS. AND BIOL. MAT.
BIOCHEMICAL REACTION KINETICS
BIOLOGICAL EFFECTS
CARBOXYLIC ACIDS
CHEMICAL COMPOSITION
COBALT
ELEMENTS
ENZYME ACTIVITY
ENZYME INHIBITORS
ENZYMES
HEME
HETEROCYCLIC ACIDS
HETEROCYCLIC COMPOUNDS
HYDRAZINE
HYDROGEN COMPOUNDS
HYDROGEN PEROXIDE
KINETICS
MANGANESE
MATHEMATICAL MODELS
METABOLISM
METALS
NITROGEN COMPOUNDS
ORGANIC ACIDS
ORGANIC COMPOUNDS
ORGANIC NITROGEN COMPOUNDS
OXIDOREDUCTASES
OXYGEN COMPOUNDS
PEROXIDASES
PEROXIDES
PIGMENTS
PORPHYRINS
PROTEINS
RADICALS
REACTION KINETICS
TRANSITION ELEMENTS