Reaction kinetics of versatile peroxidase for the degradation of lignin compounds

Abstract

The H2O2-dependent degradation of adlerol by a crude versatile peroxidase from Bjerkandera adusta, a new ligninolytic enzyme, was investigated. Adlerol (1-(3,4-dimethoxyphenyl)-2-(2-methoxyphenoxy)-1,3-propanediol)) is a non-phenolic β-O-4 dimer whose structural architecture represents the most abundant unit (50-65%) of the valuable renewable biopolymer lignin. Lignin removel plays a key role in utilizing lignocellulosic biomass in biorefineries. Steady-state analyses in the μL scale showed saturation kinetics for both, H2O2 and adlerol with quite sensitive response to H2O2. This was characterized through slow transient states (lag phases) prior steady-state and were enhanced by increasing H2O2 concentration. The major reason for such phenomena was found to be an accumulation of compound III (EIII) via reaction of compound II (EII) with H2O2; instead with adlerol to the enzyme's ground state E0 in order to restart another catalytic cycle. As result, the enzyme deviated from its normal catalytic cycle. A corresponding threshold was determined at ≥ 50 μM H2O2 and an adlerol to H2O2 ratio of 15:1 for the given conditions. Furthermore, EIII did not represent a catalytical dead-end intermediate as it is generally described. By an additional decrease of the adlerol to H2O2 ratio of ca. 3 at the latest, considerable irreversible enzyme deactivations occurred promoted through reaction of EIII with H2O2. At a mL scale deactivation kinetics by H2O2 were further examined in dependence on adlerol presence. The course followed a time-dependent irreversible deactivation (two step mechanism) and was diminished in the presence of adlerol. The deactivation could be sufficiently described by an equation similar to the Michaelis-Menten type, competitive inhibited by adlerol. Finally, first estimates of the kinetic parameters vmax, KmS1 (S1: H2O2), KmS2 (S2: adlerol), kiapp and Kiapp were made. Moreover, the peroxidase reaction mechanism was reviewed and recommendations are given preventing permature enzyme losses. © 2013 Science Publication.