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Feruloyl esterases (FAEs) and acetyl xylan esterases (AXEs) are important enzymes for plant biomass degradation and are both present in Carbohydrate Esterase family 1 (CE1) of the Carbohydrate-Active enZymes database. In this study, ten novel fungal CE1 enzymes from different subfamilies were heterologously produced and screened for their activity towards model and complex plant biomass substrates. CE1_1 enzymes possess AXE activity, while CE1_5 enzymes showed FAE activity. Two enzymes from CE1_2 and one from CE1_5 possess dual feruloyl/acetyl xylan esterase (FXE) activity, showing expansion of substrate specificity. The new FXEs from CE1 can efficiently release both feruloyl and acetyl residues from feruloylated xylan, making them particularly interesting novel components of industrial enzyme cocktails for plant biomass degradation.

Additional file 1.

Additional file 1.

Additional file 3. Structure-based amino acid “segments only” sequence alignment of AA9 LPMOs.

Additional file 4: Figure S4. Phylogenetic analysis of lignin-induced P. anserina AA7 oxidoreductases.

Additional file 2. Structure-based amino acid “full sequence” alignment of AA9 LPMOs.

Additional file 2: Proteomic analysis: Figure S2. Proteomic analysis of secreted proteins during growth of P. anserina on glucuronoarabinoxylan (GAX) and GAX + Lignin. Grouped according to functional annotations. Figure S3. Proteomic analysis of secreted individual proteins during growth of P. anserina on glucuronoarabinoxylan (GAX) and GAX + Lignin. Accession numbers according to JGI database (P. anserina mat + v1.0).

Additional file 1: Substrate characterization data: Figure S1. Py-GC-HR-MS pyrograms (TIC) of wheat straw, wheat straw lignin and glucuronoarabinoxylan (GAX). Table S1. Py-GC-HR-MS relative abundance of lignin compounds in wheat straw lignin isolate used for fermentations with P. anserina. Table S2. Semiquantitative 1H–13C HSQC NMR structural characterization of wheat straw lignin isolate used for fermentations with P. anserina.

Additional file 1. Fig. S1. MW distributions of TXG-digests from NcLPMO9C and NcLPMO9M. Fig. S2. HPAEC elution patterns of β-galactosidase treated XEG and NcLPMO9C digested TXG. Fig. S3. MALDI-TOF mass spectra of β-galactosidase treated XEG and NcLPMO9C digested TXG. Fig. S4. Negative ion mode CID-MS/MS fragmentation patterns of XXX and XLL present in the NcLPMO9C-TXG-digest. Fig. S5. Negative ion mode CID-MS/MS fragmentation patterns of XGX and LXG present in the NcLPMO9M-TXG-digest. Fig. S6. HILIC-ESI-MS base-peak chromatograms of BCXG-digests from NcLPMO9C and NcLPMO9M. Fig. S7. Phylogenetic tree in circle style of “full length” (FL) AA9 LPMOs. Fig. S8. Phylogenetic tree in circle style of “segments only” (SO) AA9 LPMOs. Fig. S9. Crystal structures and homology models of LPMOs tested on XG. Fig. S10. HPAEC elution patterns of TXG, and of TXG digested with NcLPMO9F with the addition of Asc. Fig. S11. HPAEC elution patterns of TXG, and of TXG digested with MtLPMO9I with the addition of Asc. Table S1. Selected mass (m/z) list for non-, C4- and C1-oxidized TXG oligosaccharides. Table S2. List of non-oxidized XG oligosaccharides identified based on fragmentation patterns in CID-MS/MS present in the NcLPMO9C-TXG-digest. Table S3. List of non-oxidized XG oligosaccharides identified based on fragmentation patterns in CID-MS/MS present in the NcLPMO9M-TXG-digest. Table S4. Characterized LPMOs; organism, Genbank accession number, LPMO name (if applicable), PDB entry (if applicable) and reference (if applicable).

The white button mushroom Agaricus bisporus is one of the most widely produced edible fungus with a great economical value. Its commercial cultivation process is often performed on wheat straw and animal manure based compost that mainly contains lignocellulosic material as a source of carbon and nutrients for the mushroom production. As a large portion of compost carbohydrates are left unused in the current mushroom cultivation process, the aim of this work was to study wild-type A. bisporus strains for their potential to convert the components that are poorly utilized by the commercial strain A15. We therefore focused our analysis on the stages where the fungus is producing fruiting bodies. Growth profiling was used to identify A. bisporus strains with different abilities to use plant biomass derived polysaccharides, as well as to transport and metabolize the corresponding monomeric sugars. Six wild-type isolates with diverse growth profiles were compared for mushroom production to A15 strain in semi-commercial cultivation conditions. Transcriptome and proteome analyses of the three most interesting wild-type strains and A15 indicated that the unrelated A. bisporus strains degrade and convert plant biomass polymers in a highly similar manner. This was also supported by the chemical content of the compost during the mushroom production process. Our study therefore reveals a highly conserved physiology for unrelated strains of this species during growth in compost.

Additional file 4: Figure S1. Residual lignin, glucuronoarabinoxylan (GAX) and glucan (cellulose) in fungal-treated wheat straw during growth (1, 3 and 7Â weeks). Cs Ceriporiopsis subvermispora, Pe Pleurotus eryngii, Le Lentinula edodes. Average and standard deviation of analytical triplicates on pooled biological triplicates.

Additional file 6: Figure S3. Structural changes of fungal-treated wheat straw lignin during fungal growth (1, 3, and 7 weeks) determined by semi-quantitative py-GC–MS. Cs Ceriporiopsis subvermispora, Pe Pleurotus eryngii, and Le Lentinula edodes. S/G ratio (a) and relative abundances of unsubstituted (b), Cα-oxidized (c), and Ph–Cγ (d) substructures are based on the structural classification shown in Additional file 2: Table S1. Square Cs1, filled square Cs12, triangle Pe3, filled triangle Pe6, circle Le8, filled circle Le10. Average and standard deviation of analytical triplicates on pooled biological triplicates.

Additional file 2: Table S1. Identity, structural classification and relative abundance of lignin-derived pyrolysis products by 13C-IS py-GC–MS. Control and fungal-treated wheat straw samples after 7 weeks of treatment. Cs Ceriporiopsis subvermispora, Pe Pleurotus eryngii, and Le Lentinula edodes. Average of analytical triplicates on pooled biological triplicates.

Additional file 5: Figure S2. EI–MS (70 eV) spectra and structures of vanilloyl acetaldehyde (VAL) and syringoyl acetaldehyde (SAL) observed by py-GC–MS. Compounds were identified on the basis of fragmentation pattern and retention time. Average mass spectrum across the chromatographic peak with noise subtraction at two sides.

Additional file 7: Table S3. Assignments of the lignin 13Câ 1H correlation peaks in the HSQC spectra of control and fungal-treated wheat straw. (t) Tentative assignment.

Additional file 8: Figure S4. HSQC NMR spectra of fungal-treated wheat straw. Cs12 (a, b), Pe3 (c, d), Pe6 (e, f) Le8 (g, h) Le10 (i, j). a, c, e, g, i: Aliphatic (δC/δH 50–90/2.5–6.0) region, b, d, f, h, j: aromatic/unsaturated (δC/δH 90–160/6.0–8.0) region. Structures of annotated correlation peaks are presented in Fig. 5. Gmod and Smod are presented in a lighter color of G and S, respectively. Carbohydrate and unassigned signals are presented in gray. u: unassigned signal overlapping Aα/A′α.

10% SDS-PAGE of purified endo-xylanase from G. thermodenitrificans T12 followed by PageBlue staining. Lane 1: Protein marker; Lane 2: Pellet fraction; Lane 3: cell-free extract; Lane 4: non-binding protein fraction from FPLC; Lane 5: Purified recombinant GtXynA1; Lane 6: Purified and desalted recombinant GtXynA1; Lane 7: Protein marker. (DOCX 614 kb)

Additional file 8: Table S3. Selected cellulase-rich Basidiomycota from the JGI database1.

Additional file 1: Table S1. Primers used in this study. Table S2. Statistical differences over time in MnP and Lcc activity and chitin content in compost as presented in Fig. 2. Time points sharing a letter are not significantly different (ANOVA with Bonferroni post hoc test, p

Additional file 7: Table S2. Selected cellulase-rich Ascomycota from the JGI database1.

FPLC purification of GtXynA1. Protein fraction not bound to the nickel column eluted with the first 65 mL eluent. Bound GtXynA1 protein was removed from the nickel column using a imidazole gradient (yellow line) which increased from 0 mM to 500 mM over a time span of 20 min (A). Fractions 22–25 were pooled and used for desalting the purified GtXynA1 (B). Fractions 8–10 of the desalting column were pooled and then used for further experiments. (DOCX 311 kb)

Additional file 1: Figure S1. pH (A) and temperature profiles (C) of the incubated ReGH67 with the aldouronic acids mixture (AAc) and the pH (B) and temperature profiles (D) of the incubated ReGH115 with beechwood xylan.