The characterisation of immobilised lignin peroxidase by flow injection analysis.
Immobilised lignin peroxidase has been investigated using a flow system in the steady state and by flow injection analysis (FIA). In the steady state, the extreme sensitivity of the enzyme towards inactivation by H2O2 resulted in a stable response only in the presence of saturating levels of organic substrate and at very low (10 microM) peroxide concentrations. By contrast, the low contact time during FIA led to a stable response to injections of 100 microM H2O2. At higher peroxide concentrations a reproducible inactivation was observed, allowing a study of factors affecting both activity and stability. Lignin peroxidase substrates that undergo at least semi-reversible oxidation/reduction, including high-molecular-weight lignin fractions, could be detected by electrochemical reduction of the oxidation products. With this detection system it was possible to demonstrate the role of veratryl alcohol as mediator. This mediated oxidation of lignin functioned only when all components were present simultaneously, and was not observed when lignin was separated from the site of veratryl alcohol oxidation.
Fawer
M S
MS
Institute of Biotechnology, ETH Hönggerberg, Zürich, Switzerland.
Stierli
J
J
Cliffe
S
S
Fiechter
A
A
D000097:Acetonitriles / Q000494:pharmacology; D001592:Benzyl Alcohols / Q000378:metabolism; D004126:Dimethylformamide / Q000494:pharmacology; D004146:Dioxanes / Q000494:pharmacology; D004789:Enzyme Activation; D004795:Enzyme Stability; D004800:Enzymes, Immobilized / Q000378:metabolism*; D006861:Hydrogen Peroxide / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008722:Methods; D008970:Molecular Weight; D010084:Oxidation-Reduction; D010544:Peroxidases / Q000378:metabolism*; D013379:Substrate Specificity
D000097:Acetonitriles; D001592:Benzyl Alcohols; D004146:Dioxanes; D004800:Enzymes, Immobilized; D004126:Dimethylformamide; D006861:Hydrogen Peroxide; D010544:Peroxidases; C042858:lignin peroxidase; C042197:veratryl alcohol