Microbial β-etherases, which selectively cleave the β-O-4 aryl ether linkage present in lignin, hold great promise for future applications in lignin valorization. However, very few members have been reported so far and little is known about these enzymes. By using a database mining approach, four novel bacterial β-etherases were identified, recombinantly produced in Escherichia coli, and investigated together with known β-etherases in the conversion of various lignin and non-lignin-type model compounds. The resulting activities revealed the significant influence of the substituents at the phenyl ring adjacent to the ether bond. Finally, β-etherase activity on polymeric substrates, measured by using a fluorescently labeled synthetic lignin, was also proven; this underlined the applicability of the enzymes for the conversion of lignin into renewable chemicals.

Glutathione-dependent β-etherases and glutathione lyases are key-enzymes for the biocatalytic depolymerization of lignin. In the first step, the nucleophilic attack of glutathione to the common β-O-4-arylether motif in lignin is catalyzed by β-etherases and afterwards the glutathione is removed again by the action of glutathione lyases. Given their potential impact for lignin valorization, in this paper novel glutathione lyases are reported and biocatalytically characterized based on lignin model compounds. As a result, an enzyme exhibiting increased thermostability and lowered enantioselectivitykey features for implementation of glutathione lyases in enzymatic lignin depolymerization processeswas identified. Furthermore, first mutational studies of these enzymes revealed the possibility to further alter the activity as well as enantioselectivity of glutathione lyases by means of protein engineering. From a practical perspective, onepot multi-step processes combining β-etherases and glutathione lyases are successfully set-up, giving hints on the potential that the implementation of these biocatalysts may bring for biorefinery purposes. † Electronic supplementary information (ESI) available. See

Lignin is a biomass-derived aromatic polymer that has been identified as a potential renewable source of aromatic chemicals and other valuable compounds. The valorization of lignin, however, represents a great challenge due to its high inherent functionalization, what compromises the identification of chemical routes for its selective depolymerization. In this work, an in vitro biocatalytic depolymerization process is presented, that was applied to lignin samples obtained from beech wood through OrganoCat pretreatment, resulting in a mixture of lignin-derived aromatic monomers. The reported biocracking route comprises first a laccase-mediator system to specifically oxidize the Cα hydroxyl group in the β-O-4 structure of lignin. Subsequently, selective β-O-4 ether cleavage of the oxidized β-O-4 linkages is achieved with β-etherases and a glutathione lyase. The combined enzymatic approach yielded an oily fraction of low-molecular-mass aromatic compounds, comprising coniferylaldehyde and other guaiacyl and syringyl units, as well as some larger (soluble) fractions. Upon further optimization, the reported biocatalytic route may open a valuable approach for lignin processing and valorization under mild reaction conditions.

Three new epithiodiketopiperazine natural products [outovirin A (1), outovirin B (2), and outovirin C (3)] resembling the antifungal natural product gliovirin have been identified in extracts of Penicillium raciborskii, an endophytic fungus isolated from Rhododendron tomentosum. The compounds are unusual for their class in that they possess sulfide bridges between α- and β-carbons rather than the typical α-α bridging. To our knowledge, outovirin A represents the first reported naturally produced epimonothiodiketopiperazine, and antifungal outovirin C is the first reported trisulfide gliovirin-like compound. This report describes the identification and structural elucidation of the compounds by LC-MS/MS and NMR.

Aims:  To isolate new fungal strains from subtropical soils and to identify those that produce high cellulase activity. To select microbial strains producing thermostable cellulases with potential application in industry. Methods and Results:  The new strains Penicillium sp. CR‐316 and Penicillium sp. CR‐313 have been identified and selected because they secreted a high level of cellulase in media supplemented with rice straw. Analysis by sodium dodecyl sulfate polyacrylamide gel electrophoresis, isoelectric focussing and zymography showed that the studied strains secreted multiple enzymes that hydrolyse cellulose. Cellulase activity of Penicillium sp. CR‐316, the strain showing higher production, was analysed. Optimum temperature and pH of carboxymethyl cellulase activity were 65°C and pH 4·5, respectively. Activity remained stable after incubation at 60°C and pH 4·5 for 3 h. Conclusions:  Fungal strains that secrete high levels of cellulase activity have been characterized and selected from soil. The isolated strains have complex sets of enzymes for cellulose degradation. Crude cellulase produced by Penicillium sp. CR‐316 showed activity and stability at high temperature. Significance and Impact of the Study:  Two fungal strains with biotechnological potential have been isolated. The strains secrete high levels of cellulase, and one of them, Penicillium sp. CR‐316, produces a thermostable cellulase, that makes it a good candidate for industrial applications.