Lytic polysaccharide monooxygenases utilise reducing agents within the biomass substrate to act synergistically with canonical hydrolases to enhance cellulose deconstruction.

BackgroundCurrently, the amount of protein/enzyme required to achieve effective cellulose hydrolysis is still too high. One way to reduce the amount of protein/enzyme required is to formulate a more efficient enzyme cocktail by adding so-called accessory enzymes such as xylanase, lytic polysaccharide monooxygenase (AA9, formerly known as GH61), etc., to the cellulase mixture. Previous work has shown the strong synergism that can occur between cellulase and xylanase mixtures during the hydrolysis of steam pretreated corn stover, requiring lower protein loading to achieve effective hydrolysis. However, relatively high loadings of xylanases were required. When family 10 and 11 endo-xylanases and family 5 xyloglucanase were supplemented to a commercial cellulase mixture varying degrees of improved hydrolysis over a range of pretreated, lignocellulosic substrates were observed.ResultsThe potential synergistic interactions between cellulase monocomponents and hemicellulases from family 10 and 11 endo-xylanases (GH10 EX and GH11 EX) and family 5 xyloglucanase (GH5 XG), during hydrolysis of various steam pretreated lignocellulosic substrates, were assessed. It was apparent that the hydrolytic activity of cellulase monocomponents was enhanced by the addition of accessory enzymes although the “boosting” effect was highly substrate specific. The GH10 EX and GH5 XG both exhibited broad substrate specificity and showed strong synergistic interaction with the cellulases when added individually. The GH10 EX was more effective on steam pretreated agriculture residues and hardwood substrates whereas GH5 XG addition was more effective on softwood substrates. The synergistic interaction between GH10 EX and GH5 XG when added together further enhanced the hydrolytic activity of the cellulase enzymes over a range of pretreated lignocellulosic substrates. GH10 EX addition could also stimulate further cellulose hydrolysis when added to the hydrolysis reactions when the rate of hydrolysis had levelled off.ConclusionsEndo-xylanases and xyloglucanases interacted synergistically with cellulases to improve the hydrolysis of a range of pretreated lignocellulosic substrates. However, the extent of improved hydrolysis was highly substrate dependent. It appears that those accessory enzymes, such as GH10 EX and GH5 XG, with broader substrate specificities promoted the greatest improvements in the hydrolytic performance of the cellulase mixture on all of the pretreated biomass substrates.

Some of the most damaging tree pathogens can attack woody stems, causing lesions (cankers) that may be lethal. To identify the genomic determinants of wood colonization leading to canker formation, we sequenced the genomes of the poplar canker pathogen, Mycosphaerella populorum, and the closely related poplar leaf pathogen, M. populicola. A secondary metabolite cluster unique to M. populorum is fully activated following induction by poplar wood and leaves. In addition, genes encoding hemicellulosedegrading enzymes, peptidases, and metabolite transporters were more abundant and were up-regulated in M. populorum growing on poplar wood-chip medium compared with M. populicola. The secondary gene cluster and several of the carbohydrate degradation genes have the signature of horizontal transfer from ascomycete fungi associated with wood decay and from prokaryotes. Acquisition and maintenance of the gene battery necessary for growth in woody tissues and gene dosage resulting in gene expression reconfiguration appear to be responsible for the adaptation of M. populorum to infect, colonize, and cause mortality on poplar woody stems. poplar pathogen | tree disease | fungal genomics | Septoria canker

No abstract

Effective enzymatic hydrolysis of insoluble cellulose requires the synergistic action of a suite of cellulase components. Most previous studies have only assessed cellulase synergism on model cellulosic substrates. When the actions of individual and combinations of cellulases (Cel7A, Cel6A, Cel7B, Cel5A) were assessed on various pretreated lignocellulosic substrates, Cel7A was shown to be the major contributor to overall cellulose hydrolysis, with the other enzymes synergistically enhancing its hydrolytic efficiency, at least partially, by facilitating Cel7A desorption (assessed by a double-sandwich enzyme-linked immunosorbent assay). When the influences of various substrate physicochemical characteristics on the effectiveness of enzyme synergism were assessed, a strong relationship was observed between cellulose accessibility (as determined by the cellulose binding module technique) and the degree of synergism, with greater synergy observed on the more disorganized/accessible cellulose surface.