Substrate specificity of lignin peroxidase and a S168W variant of manganese peroxidase

Area of Science:

• Biochemistry
• Enzymology
• Fungal Metabolism

Background:

• Lignin peroxidase (LiP) and manganese peroxidase (MnP) are heme-containing enzymes from white-rot fungi.
• MnP exhibits specificity for Mn(2+), while LiP has broad substrate specificity.
• The structural basis for LiP's broad substrate specificity remains unclear.

Purpose of the Study:

• To investigate the substrate specificity of a S168W variant of Phanerochaete chrysosporium MnP.
• To determine if a single oxidation site can account for broad substrate specificity.

Main Methods:

• Site-directed mutagenesis of Phanerochaete chrysosporium MnP to create the S168W variant.
• Enzymatic assays measuring the oxidation of various small molecule and polymeric substrates.
• Kinetic analysis and stoichiometric determination of substrate oxidation.

Main Results:

• The S168W MnP variant retained full Mn(2+) oxidase activity.
• The S168W MnP variant oxidized a wide range of LiP substrates, unlike native MnP.
• Kinetic and stoichiometric data for veratryl alcohol oxidation were similar between the MnP variant and LiP.
• Specific and nonspecific oxidation patterns for guaiacol and ferrocyanide were observed for the MnP variant and LiP.

Conclusions:

• The S168W MnP variant functionally mimics LiP, oxidizing diverse substrates.
• These findings support the hypothesis that a single oxidation site near the surface tryptophan confers broad substrate specificity.
• This study provides insights into the catalytic mechanisms of lignin-degrading enzymes.