Database Mining for Novel Bacterial ß-Etherases, Glutathione-Dependent Lignin-Degrading Enzymes.

Lignin is the most abundant aromatic polymer in nature and a promising renewable source for the provision of aromatic platform chemicals and biofuels. ß-Etherases are enzymes with a promising potential for application in lignin depolymerization due to their selectivity in the cleavage of ß-O-4 aryl ether bonds. However, only a very limited number of these enzymes have been described and characterized so far. Using peptide pattern recognition (PPR) as well as phylogenetic analyses, 96 putatively novel ß-etherases have been identified, some even originating from bacteria outside the order Sphingomonadales A set of 13 diverse enzymes was selected for biochemical characterization, and ß-etherase activity was confirmed for all of them. Some enzymes displayed up to 3-fold higher activity than previously known ß-etherases. Moreover, conserved sequence motifs specific for either LigE- or LigF-type enzymes were deduced from multiple-sequence alignments and the PPR-derived peptides. In combination with structural information available for the ß-etherases LigE and LigF, insight into the potential structural and/or functional role of conserved residues within these sequence motifs is provided. Phylogenetic analyses further suggest the presence of additional bacterial enzymes with potential ß-etherase activity outside the classical LigE- and LigF-type enzymes as well as the recently described heterodimeric ß-etherases.IMPORTANCE The use of biomass as a renewable source and replacement for crude oil for the provision of chemicals and fuels is of major importance for current and future societies. Lignin, the most abundant aromatic polymer in nature, holds promise as a renewable starting material for the generation of required aromatic structures. However, a controlled and selective lignin depolymerization to yield desired aromatic structures is a very challenging task. In this regard, bacterial ß-etherases are especially interesting, as they are able to cleave the most abundant bond type in lignin with high selectivity. With this study, we significantly expanded the toolbox of available ß-etherases for application in lignin depolymerization and discovered more active as well as diverse enzymes than previously known. Moreover, the identification of further ß-etherases by sequence database mining in the future will be facilitated considerably through our deduced etherase-specific sequence motifs.

Microbial ß-etherases and glutathione lyases for lignin valorisation in biorefineries: current state and future perspectives.

Lignin is the major aromatic biopolymer in nature, and it is considered a valuable feedstock for the future supply of aromatics. Hence, its valorisation in biorefineries is of high importance, and various chemical and enzymatic approaches for lignin depolymerisation have been reported. Among the enzymes known to act on lignin, ß-etherases offer the possibility for a selective cleavage of the ß-O-4 aryl ether bonds present in lignin. These enzymes, together with glutathione lyases, catalyse a reductive, glutathione-dependent ether bond cleavage displaying high stereospecificity. ß-Etherases and glutathione lyases both belong to the superfamily of glutathione transferases, and several structures have been solved recently. Additionally, different approaches for their application in lignin valorisation have been reported in the last years. This review gives an overview on the current knowledge on ß-etherases and glutathione lyases, their biochemical and structural features, and critically discusses their potential for application in biorefineries.