Haloperoxidase activity of Phanerochaete chrysosporium lignin peroxidases H2 and H8.
Monochlorodimedone (MCD), commonly used as a halogen acceptor for haloperoxidase assays, was oxidized by hydrogen peroxide in the presence of lignin peroxidase isoenzymes H2 and H8. When oxidized, it produced a weak absorption band with an intensity that varied with pH. This absorbance was used as a simple method for the product analysis because it disappeared when MCD was brominated or chlorinated. We assessed the activity of the lignin peroxidases for oxidation of bromide by measuring the bromination of MCD, the formation of tribromide, the bromide-mediated oxidation of glutathione, and the bromide-mediated catalase-like activity. We analyzed the reaction products of MCD and the halide-mediated oxidation of glutathione when bromide was replaced by chloride. These enzymes demonstrated no significant activity for oxidation of chloride. Unlike other peroxidases, the lignin peroxidases exhibited similar pH-activity curves for the iodide and bromide oxidations. The optimum pH for activity was about 2.5. Surprisingly, this pH dependence of lignin peroxidase activity for the halides was nearly the same in the reactions with hydrogen donors, such as hydroquinone and guaiacol. The results suggested that protonation of the enzymes with pKa approximately 3.2 is necessary for the catalytic function of lignin peroxidases, irrespective of whether the substrates are electron or hydrogen donors. These unique reaction profiles of lignin peroxidases are compared to those of other peroxidases, such as lactoperoxidase, bromoperoxidase, chloroperoxidase, and horseradish peroxidase. Isozyme H2 was more active than isozyme H8, but isozyme H8 was more stable at very acidic pH.
Farhangrazi
Z S
ZS
National Center for the Design of Molecular Function, Utah State University, Logan 84322-4630.
Sinclair
R
R
Yamazaki
I
I
Powers
L S
LS
D001487:Basidiomycota / Q000201:enzymology*; D001965:Bromides / Q000378:metabolism; D002712:Chlorides / Q000378:metabolism; D003512:Cyclohexanones / Q000378:metabolism; D005978:Glutathione / Q000378:metabolism; D006861:Hydrogen Peroxide / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007454:Iodides / Q000378:metabolism; D007527:Isoenzymes / Q000378:metabolism*; D007700:Kinetics; D010084:Oxidation-Reduction; D010544:Peroxidases / Q000378:metabolism*; D013053:Spectrophotometry
D001965:Bromides; D002712:Chlorides; D003512:Cyclohexanones; D007454:Iodides; D007527:Isoenzymes; C006991:chlorodimedone; D006861:Hydrogen Peroxide; D010544:Peroxidases; C042858:lignin peroxidase; D005978:Glutathione