Agaricales Mushroom Lignin Peroxidase: From Structure–Function to Degradative Capabilities

Abstract

Lignin biodegradation has been extensively studied in white-rot fungi, which largely belong to order Polyporales. Among the enzymes that wood-rotting polypores secrete, lignin peroxidases (LiPs) have been labeled as the most efficient. Here, we characterize a similar enzyme (ApeLiP) from a fungus of the order Agaricales (with ~13,000 described species), the soil-inhabiting mushroom Agrocybe pediades. X-ray crystallography revealed that ApeLiP is structurally related to Polyporales LiPs, with a conserved heme-pocket and a solvent-exposed tryptophan. Its biochemical characterization shows that ApeLiP can oxidize both phenolic and non-phenolic lignin model-compounds, as well as different dyes. Moreover, using stopped-flow rapid spectrophotometry and 2D-NMR, we demonstrate that ApeLiP can also act on real lignin. Characterization of a variant lacking the above tryptophan residue shows that this is the oxidation site for lignin and other high redox-potential substrates, and also plays a role in phenolic substrate oxidation. The reduction potentials of the catalytic-cycle intermediates were estimated by stopped-flow in equilibrium reactions, showing similar activation by H2O2, but a lower potential for the rate-limiting step (compound-II reduction) compared to other LiPs. Unexpectedly, ApeLiP was stable from acidic to basic pH, a relevant feature for application considering its different optima for oxidation of phenolic and nonphenolic compounds.

Visible light-exposed lignin facilitates cellulose solubilization by lytic polysaccharide monooxygenases

“…We show here that LPMOs can oxidize polymeric lignin directly to recruit electrons and do so at an appreciable rate. The rates determined in our stopped-flow experiments are one order of magnitude lower than those observed for lignin oxidation by manganese peroxidase 60 , between two and three orders of magnitude lower than the most efficient lignin peroxidases 61 , and two orders of magnitude lower than LPMO reduction by one of the most efficient small molecule reductants, AscA 12 .…”

Evaluation of the lignocellulose degradation potential of Mediterranean forests soil microbial communities through diversity and targeted functional metagenomics

“…Further, the recovery of MnPs from several different genera of Russulales, Hymenochaetales, Agaricales, and Polyporales, irrespective of their abundance in soil samples, strongly indicates that the set of primers used in this work in nested PCR is highly efficient for the recovery of AA2 enzymes from all fungi possessing such genes. Although it was previously thought that Agaricales lack LiP with the characteristic amino acid residues of the white-rot Polyporales, such molecule was recently detected in Agrocybe, albeit of the lower enzymatic activity due to the differentiation of the neighboring residues (Sánchez-Ruiz et al, 2021). This novel finding combined with the numerous peroxidase sequences recorded in this work, show that there is still plenty to discover about Class II peroxidases, and that genomic, metagenomic and transcriptomic approaches will certainly pave the way to uncover novel lignin degrading enzymes in the quest of effective renewable energy solutions.…”

Multicopper oxidases with laccase-ferroxidase activity: Classification and study of ferroxidase activity determinants in a member from Heterobasidion annosum s. l.

“…Stock solutions were routinely prepared in 0–100 % ethanol and Milli-Q H 2 O. All reactions were performed in the same fashion excepting for Fe (II) (described in Section 2.6 ) and Mn (II) oxidation [47] . Reactions in triplicate were carried out in 96-well plates (UV-Star 96-well plates when needed) by adding 20 µL of pure diluted enzyme to a mixture reaction of 160 µL of CP buffer (final concentration in well: 100 mM) at pH 4 for azo dyes and aromatic amines, and pH 5 for phenols and Mn (II).…”