| Various fungi secrete laccase to degrade lignin for source of carbon;however,the purified laccase presents extremely low catalytic activity,for lack of proper mediators,such as ABTS,HBT and TEMPO.These mediators,can mediate the oxidation reaction on lignin and promote laccase’s catalytic ability.Nevertheless,their high environmental costs and inclination to produce toxic products hamper them from further application on an industrial scale.Due to the aforementioned shortcomings of laccase artificial mediators,it has now become one of the hot spots to find stable,eco-friendly and low toxicity mediators in the research field of laccase environmental protection and biomass energy application..Here we employed extraction and column chromatography on Pleurotus eryngii,measured regulation degree(RD%)for each component with fast LMS(laccase-mediator-system)enzyme kinetics experiment,and analyzed the potential mediator molecules that were enriched in the most active components with the mummichog algorithm implemented in XCMS metabolomics package.It was found that toluene-extract of compost was the most and further comparative analysis on its most active component CET05 obtained the most significant metabolites,which are involved in six metabolic pathways,i.e.,suberin monomers biosynthesis,coumarins biosynthesis,eugenol and isoeugenol biosynthesis et al.Moreover,these natural mediator molecules from the most active component exhibited physicochemical similarity.Besides study on small molecule mediators,we also conducted further research on the interaction mode between heme proteins and laccase.It has been recently acquainted that laccase together with proteins containing heme might produce a synergistic effect in a complicated ligninase system in fungi.However,it was yet unknown how those metalloproteins work cooperatively in lignin biodegradation.Here we carried out a mechanistic investigation on hemoglobin-mediated laccase oxidation catalysis on lignosulfonates as a soluble lignin substrate.With a combination of UVvisible spectroscopy,isothermal titration calorimetry(ITC),gel permeation chromatography(GPC),Raman spectroscopy,2D-NMR,and HPLC-MS,two comparative systems with the absence and presence of hemoglobin were characterized for the laccase-catalyzed reaction.It was found that hemoglobin not only physically interacted with the substrate,but also chemically regulated the oxidative product profile.It was suggested that hemoglobin was involved in the process of protein complexation,substrate absorption,oxygen transportation,and peroxide dismutation.Finally,degradation rate of the laccase and laccase-hemoglobin system on phenolic and aniline compounds was compared,and the feasibility of the using laccase-hemoglobin system in the treatment of pollutants was confirmed. |
