Detection and characterization of the lignin peroxidase compound II-veratryl alcohol cation radical complex.
Lignin peroxidases (LiP) from the white-rot fungus Phanerochaete chrysosporium oxidize veratryl alcohol (VA) by two electrons to veratryl aldehyde, although the VA cation radical (VA.+) is an intermediate [Khindaria, A., et al. (1995) Biochemistry 34, 6020-6025]. It was speculated, on the basis of kinetic evidence, that VA*+ can form a catalytic complex with LiP compound II. We have used low-temperature EPR to provide direct evidence for the formation of the complex. The EPR spectrum of VA*+ obtained at 4 K was explained by a model for coupling between the oxoferryl moiety of the heme (S = 1) and VA.+ (S = 1/2) similar to the model proposed for an oxyferryl and a porphyrin pi cation radical of horseradish peroxidase. The coupling constant suggested that VA.+ was equally ferro- and antiferromagnetically coupled to the oxoferryl moiety. The spectrum was simulated with g perpendicular only marginally greater than g parallel. This was surprising since the only other known organic radical coupled to the heme iron in a peroxidase is the tryptophan cation radical in cytochrome c peroxidase which exhibits a g tensor with g parallel greater than g perpendicular. Spin concentration analysis suggested that the 1 mol of VA*+ was coupled to the oxoferryl moiety per mole of enzyme. The VA.+ signal decayed with a first-order decay constant of 1.76 s-1, in close agreement with the earlier published decay constant of 1.85 s-1 from room-temperature EPR studies. The exchange coupling between VA.+ and the oxoferryl moiety strongly advocates calling this species (VA.+ and LiP compound II) a catalytic complex.
Khindaria
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Biotechnology Center, Utah State University, Logan, Utah 84322-4705, USA.
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Aust
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D001487:Basidiomycota / Q000201:enzymology*; D001592:Benzyl Alcohols / Q000378:metabolism*; D002412:Cations; D003080:Cold Temperature; D004578:Electron Spin Resonance Spectroscopy; D005609:Free Radicals; D006420:Hemeproteins / Q000378:metabolism*; D007527:Isoenzymes / Q000378:metabolism*; D010544:Peroxidases / Q000378:metabolism*
D001592:Benzyl Alcohols; D002412:Cations; D005609:Free Radicals; D006420:Hemeproteins; D007527:Isoenzymes; D010544:Peroxidases; C042858:lignin peroxidase; C042197:veratryl alcohol