CAZyme prediction in ascomycetous yeast genomes guides discovery of novel xylanolytic species with diverse capacities for hemicellulose hydrolysis

Abstract

Background Ascomycetous yeasts from the kingdom fungi inhabit every biome in nature. While filamentous fungi have been studied extensively regarding their enzymatic degradation of the complex polymers comprising lignocellulose, yeasts have been largely overlooked. As yeasts are key organisms used in industry, understanding their enzymatic strategies for biomass conversion is an important factor in developing new and more efficient cell factories. The aim of this study was to identify polysaccharide-degrading yeasts by mining CAZymes in 332 yeast genomes from the phylum Ascomycota. Selected CAZyme-rich yeasts were then characterized in more detail through growth and enzymatic activity assays. Results The CAZyme analysis revealed a large spread in the number of CAZyme-encoding genes in the ascomycetous yeast genomes. We identified a total of 217 predicted CAZyme families, including several CAZymes likely involved in degradation of plant polysaccharides. Growth characterization of 40 CAZyme-rich yeasts revealed no cellulolytic yeasts, but several species from the Trichomonascaceae and CUG-Ser1 clades were able to grow on xylan, mixed-linkage β-glucan and xyloglucan. Blastobotrys mokoenaii, Sugiyamaella lignohabitans, Spencermartinsiella europaea and several Scheffersomyces species displayed superior growth on xylan and well as high enzymatic activities. These species possess genes for several putative xylanolytic enzymes, including ones from the well-studied xylanase-containing glycoside hydrolase families GH10 and GH30, which appear to be attached to the cell surface. B. mokoenaii was the only species containing a GH11 xylanase, which was shown to be secreted. Surprisingly, no known xylanases were predicted in the xylanolytic species Wickerhamomyces canadensis, suggesting that this yeast possesses novel xylanases. In addition, by examining non-sequenced yeasts closely related to the xylanolytic yeasts, we were able to identify novel species with high xylanolytic capacities. Conclusions Our approach of combining high-throughput bioinformatic CAZyme-prediction with growth and enzyme characterization proved to be a powerful pipeline for discovery of novel xylan-degrading yeasts and enzymes. The identified yeasts display diverse profiles in terms of growth, enzymatic activities and xylan substrate preferences, pointing towards different strategies for degradation and utilization of xylan. Together, the results provide novel insights into how yeast degrade xylan, which can be used to improve cell factory design and industrial bioconversion processes.

Oral and Gut Microbial Carbohydrate-Active Enzymes Landscape in Health and Disease

“…Characterizing the CAZyme complement encoded by the microbial genetic diversity present in an individual host (CAZy-typing) can be useful in predicting carbohydrate pools that the host can metabolize, or which CAZyme families are underrepresented requiring supplementation via microbiota transplantation or probiotics. The usefulness of such predictions is increasingly becoming relevant in clear hypotheses formulation and guided experimentation as has recently been applied in identifying hemicellulose hydrolytic yeast species (Ravn et al, 2021). In order to link disease state with CAZyme profiles, we used publicly available metagenomes to compare the abundance and distribution of CAZymes in saliva and stool samples from healthy subjects and patients suffering from colorectal cancer, rheumatoid arthritis, and type 1 diabetes.…”

Yeast GH30 Xylanase from Sugiyamaella lignohabitans Is a Glucuronoxylanase with Auxiliary Xylobiohydrolase Activity

“…Compared to bacterial and fungal xylanases, yeast xylanases have not been studied so extensively, presumably due to much lower levels of xylanase production by the yeasts. The best characterized yeast xylanases are from yeast-like fungus Aureobasidium pullulans [ 16 , 17 ] and from several Cryptococcus species [ 18 , 19 , 20 ], but EXs were purified and characterized also from Pichia ( Scheffersomyces ), Pseudozyma and Blastobotrys genera [ 21 , 22 , 23 ]. All of them belong to either the GH10 or GH11 family.…”

“…Bioinformatic mining in 332 yeast genomes from the phylum Ascomycota has revealed only a few putative yeast xylanases: one from the GH11 family ( Blastobotrys mokoenii ), five from the GH10 family (two from Spencermartinsiella europaea, two from Sugiyamaella lignohabitans and one from B. peoriensis ) and three from the GH30-7 subfamily ( Sp. europaea, Su lignohabitans, B. mokoenii ) [ 23 ].…”

Development of a Vector Set for High or Inducible Gene Expression and Protein Secretion in the Yeast Genus Blastobotrys

“…In 2021, Ravn et al looked for polysaccharide-hydrolysing enzymes in 332 yeast genomes from the Ascomycota phylum and identified several new xylan-degrading species from the Trichomonascaceae family. Notably, they identified surface-anchored xylanases of the GH10 family in several species and reported the presence of a secreted xylanase of the GH11 family in Blastobotrys mokoenaii [ 37 ]. They highlighted that the eight species from the Trichomonascaceae family have a more diverse and abundant xylanolytic CAZyme distribution than yeasts from other families and classified B. mokoenaii as the best xylanolytic yeast.…”