Abstract:Lignin is 1 of the 3 major components of lignocellulose. Its polymeric structure includes aromatic subunits that can be converted into high-value-added products, but this potential cannot yet been fully exploited because lignin is highly recalcitrant to degradation. Different approaches for the depolymerization of lignin have been tested, including pyrolysis, chemical oxidation, and hydrolysis under supercritical conditions. An additional strategy is the use of lignin-degrading enzymes, which imitates the natu… Show more
“…Many research groups use E. coli as an expression host, but the peroxidases mostly aggregate in inclusion bodies and have to be tediously refolded . Peroxidase expression levels in heterologous hosts have not yet exceeded 100 mg L −1 in the culture medium, but are mostly in the range of 1–20 mg L −1 . Thus, the concentration of 132 mg L −1 achieved in this work is, to the best of our knowledge, the highest yield of ligninolytic peroxidase obtained so far.…”
Manganese peroxidases, lignin peroxidases, and versatile peroxidases secreted by white rot fungi are supposed to play an essential role in lignin degradation. Thus, these enzymes have attracted significant attention as potential biocatalysts. Versatile peroxidases are the most interesting ones, since they comprise activities of manganese and lignin peroxidases. Herein, we demonstrate how the properties of a new manganese peroxidase from Moniliophthora roreri, designated MrMnP1, were shifted towards those of a versatile peroxidase. MrMnP1 was cloned in Pichia pastoris X‐33 and highly expressed in a fed‐batch fermentation, yielding 132 mg L−1 of active enzyme. To extend the substrate spectrum of MrMnP1, a catalytically active tryptophan present in lignin and versatile peroxidases was first introduced. Additionally, the role of five amino acids at positions adjacent to the catalytic tryptophan was elucidated through their replacement by those found in a versatile peroxidase from Pleurotus eryngii. The resulting mutants demonstrated new activities towards high‐redox‐potential substrates, such as lignin dimers, veratryl alcohol, and the azo dye Reactive Black 5.
“…Many research groups use E. coli as an expression host, but the peroxidases mostly aggregate in inclusion bodies and have to be tediously refolded . Peroxidase expression levels in heterologous hosts have not yet exceeded 100 mg L −1 in the culture medium, but are mostly in the range of 1–20 mg L −1 . Thus, the concentration of 132 mg L −1 achieved in this work is, to the best of our knowledge, the highest yield of ligninolytic peroxidase obtained so far.…”
Manganese peroxidases, lignin peroxidases, and versatile peroxidases secreted by white rot fungi are supposed to play an essential role in lignin degradation. Thus, these enzymes have attracted significant attention as potential biocatalysts. Versatile peroxidases are the most interesting ones, since they comprise activities of manganese and lignin peroxidases. Herein, we demonstrate how the properties of a new manganese peroxidase from Moniliophthora roreri, designated MrMnP1, were shifted towards those of a versatile peroxidase. MrMnP1 was cloned in Pichia pastoris X‐33 and highly expressed in a fed‐batch fermentation, yielding 132 mg L−1 of active enzyme. To extend the substrate spectrum of MrMnP1, a catalytically active tryptophan present in lignin and versatile peroxidases was first introduced. Additionally, the role of five amino acids at positions adjacent to the catalytic tryptophan was elucidated through their replacement by those found in a versatile peroxidase from Pleurotus eryngii. The resulting mutants demonstrated new activities towards high‐redox‐potential substrates, such as lignin dimers, veratryl alcohol, and the azo dye Reactive Black 5.
“…Lactobacillus and Clostridium species are typical lactic acid-producing and acetate-oxidizing bacteria and are often reported as the dominant consumers of soluble and easily degradable organic matter (such as lipid, protein and starch) present at the onset of composting [ 54 , 55 ]. By contrast, thermophilic Saccharomonospora species predominated during the later phases of composting, presumably as they are able to degrade recalcitrant compounds such as xylan and lignin [ 56 , 57 ]. Fig.…”
BackgroundAnimal manure is a reservoir of antibiotic resistance genes (ARGs) that pose a potential health risk globally, especially for resistance to the antibiotics commonly used in livestock production (such as tetracycline, sulfonamide, and fluoroquinolone). Currently, the effects of biological treatment (composting) on the transcriptional response of manure ARGs and their microbial hosts are not well characterized. Composting is a dynamic process that consists of four distinct phases that are distinguished by the temperature resulting from microbial activity, namely the mesophilic, thermophilic, cooling, and maturing phases. In this study, changes of resistome expression were determined and related to active microbiome profiles during the dynamic composting process. This was achieved by integrating metagenomic and time series metatranscriptomic data for the evolving microbial community during composting.ResultsComposting noticeably reduced the aggregated expression level of the manure resistome, which primarily consisted of genes encoding for tetracycline, vancomycin, fluoroquinolone, beta-lactam, and aminoglycoside resistance, as well as efflux pumps. Furthermore, a varied transcriptional response of resistome to composting at the ARG levels was highlighted. The expression of tetracycline resistance genes (tetM-tetW-tetO-tetS) decreased during composting, where distinctive shifts in the four phases of composting were related to variations in antibiotic concentration. Composting had no effect on the expression of sulfonamide and fluoroquinolone resistance genes, which increased slightly during the thermophilic phase and then decreased to initial levels. As indigenous populations switched greatly throughout the dynamic composting, the core resistome persisted and their reservoir hosts’ composition was significantly correlated with dynamic active microbial phylogenetic structure. Hosts for sulfonamide and fuoroquinolone resistance genes changed notably in phylognetic structure and underwent an initial increase and then a decrease in abundance. By contrast, hosts for tetracycline resistance genes (tetM-tetW-tetO-tetS) exhibited a constant decline through time.ConclusionsThe transcriptional patterns of a core resistome over the course of composting were identified, and microbial phylogeny was the key determinant in defining the varied transcriptional response of resistome to this dynamic biological process. This research demonstrated the benefits of composting for manure treatment. It reduced the risk of emerging environmental contaminants such as tetracyclines, tetracycline resistance genes, and clinically relevant pathogens carrying ARGs, as well as RNA viruses and bacteriophages.Electronic supplementary materialThe online version of this article (doi:10.1186/s40168-017-0324-0) contains supplementary material, which is available to authorized users.
“…Ligninolytic enzyme cocktails have potential for enzymatic pretreatment of lignocellulose (Schroyen et al, 2015) as well as generating lignin-derived precursors of aromatic building blocks (Abdelaziz et al, 2016;Lubbers et al, 2019). Heterologous production of fungal ligninolytic cocktails is challenging because of the low yields of catalytically active lignin modifying peroxidases (Lambertz et al, 2016). Therefore, induction of ligninolytic enzymes in a natural system, such as a wood rotting fungus, is an attractive alternative.…”
Heterologous production of fungal ligninolytic cocktails is challenging due to the low yields of catalytically active lignin modifying peroxidases. Production using a natural system, such as a wood-rotting fungus, is a promising alternative if specific or preferential induction of the ligninolytic activities could be achieved. Using transcriptomics, gene expression of the white-rot Dichomitus squalens during growth on mixtures of aromatic compounds, with ring structures representing the two major lignin sub-units, was compared to a wood substrate. Most of the genes encoding lignin modifying enzymes (laccases and peroxidases) categorised as highly or moderately expressed on wood were expressed similarly on aromatic compounds. Higher expression levels of a subset of manganese and versatile peroxidases was observed on di-compared to mono-methoxylated aromatics. The expression of polysaccharide degrading enzymes was lower on aromatic compounds compared to wood, demonstrating that the induction of lignin modifying enzymes became more specific. This study suggests potential for aromatic waste streams, e.g. from lignocellulose pretreatment, to produce a lignin-specific enzyme cocktail from D. squalens or other white-rot fungi.
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