The importance of sourcing enzymes from non-conventional fungi for metabolic engineering and biomass breakdown

Abstract

A wealth of fungal enzymes has been identified from nature, which continue to drive strain engineering and bioprocessing for a range of industries. However, while a number of clades have been investigated, the vast majority of the fungal kingdom remains unexplored for industrial applications. Here, we discuss selected classes of fungal enzymes that are currently in biotechnological use, and explore more basal, non-conventional fungi and their underexploited biomass-degrading mechanisms as promising agents in the transition towards a bio-based society. Of special interest are anaerobic fungi like the Neocallimastigomycota, which were recently found to harbor the largest diversity of biomass-degrading enzymes among the fungal kingdom. Enzymes sourced from these basal fungi have been used to metabolically engineer substrate utilization in yeast, and may offer new paths to lignin breakdown and tunneled biocatalysis. We also contrast classic enzymology approaches with emerging 'omics'-based tools to decipher function within novel fungal isolates and identify new promising enzymes. Recent developments in genome editing are expected to accelerate discovery and metabolic engineering within these systems, yet are still limited by a lack of high-resolution genomes, gene regulatory regions, and even appropriate culture conditions. Finally, we present new opportunities to harness the biomass-degrading potential of undercharacterized fungi via heterologous expression and engineered microbial consortia.

Biomass‐degrading enzymes are catabolite repressed in anaerobic gut fungi

“…To combat this limitation, current industrial practices for biofuel and bio‐based chemical production incorporate extensive pretreatment via incubation with dilute acid or ammonium fiber explosion in order to reduce lignin content and increase accessibility to cellulose . Commercially purified cellulases and saccharolytic enzymes are subsequently added to the pretreated mixture, generally limited to enzymes from Trichoderma and Aspergillus species, which have been used largely because these aerobic fungi secrete large volumes of enzymes and are ubiquitous and easy to culture …”

Proteome specialization of anaerobic fungi during ruminal degradation of recalcitrant plant fiber

“…Enumerating anaerobic rumen fungi is challenging, mainly due to their different life stages and their growth within plant fragments as well as sub-optimal DNA extraction and molecular methods to recover their genomic information [11][12][13] . Reported counts of fungal cells vary greatly between studies, with numbers ranging between 10 3 and 10 6 cells/ml of rumen fluid [14][15][16] .…”

“…The importance of rumen fungi for biomass degradation has since then been supported by in vivo studies [23][24][25] , and recently reinforced in transcriptome studies revealing that the fungi express a range of CAZymes when grown on different carbon sources 9,26 . Although enzymes of fungal origin have been regularly explored for their remarkable capacity to degrade lignocellulosic fiber 12,27,28 , their functional role in native anaerobic habitats and within the biomass-degrading enzyme repertoire of the rumen microbiome remains unclear. Thus, we lack a complete understanding of their biology and their contribution to the function and health of the rumen ecosystems.…”

In Silico Identification of Microbial Partners to Form Consortia with Anaerobic Fungi

“…This could necessitate the addition of (expensive) beta-glucosidases, to convert cellobiose to glucose, in some applications. It is hypothesized that under-explored fungal clades, like Neocallimastigomycota, coud offer substantial benefits in this regard [11].…”

“…Given the wealth of omics-related data available, we speculate that model driven design could elucidate some of these questions [11]. Indeed, model driven analysis has successfully been used to study anaerobic organisms [16].…”