Abstract
BackgroundSugarcane bagasse has been proposed as a lignocellulosic residue for second-generation ethanol (2G) produced by breaking down biomass into fermentable sugars. The enzymatic cocktails for biomass degradation are mostly produced by fungi, but low cost and high efficiency can consolidate 2G technologies. A. fumigatus plays an important role in plant biomass degradation capabilities and recycling. To gain more insight into the divergence in gene expression during steam-exploded bagasse (SEB) breakdown, this study profiled the transcriptome of A. fumigatus by RNA sequencing to compare transcriptional profiles of A. fumigatus grown on media containing SEB or fructose as the sole carbon source. Secretome analysis was also performed using SDS-PAGE and LC-MS/MS.ResultsThe maximum activities of cellulases (0.032 U mL-1), endo-1,4-β--xylanase (10.82 U mL-1) and endo-1,3-β glucanases (0.77 U mL-1) showed that functional CAZymes (carbohydrate-active enzymes) were secreted in the SEB culture conditions. Correlations between transcriptome and secretome data identified several CAZymes in A. fumigatus. Particular attention was given to CAZymes related to lignocellulose degradation and sugar transporters. Genes encoding glycoside hydrolase classes commonly expressed during the breakdown of cellulose, such as GH-5, 6, 7, 43, 45, and hemicellulose, such as GH-2, 10, 11, 30, 43, were found to be highly expressed in SEB conditions. Lytic polysaccharide monooxygenases (LPMO) classified as auxiliary activity families AA9 (GH61), CE (1, 4, 8, 15, 16), PL (1, 3, 4, 20) and GT (1, 2, 4, 8, 20, 35, 48) were also differentially expressed in this condition. Similarly, the most important enzymes related to biomass degradation, including endoxylanases, xyloglucanases, β-xylosidases, LPMOs, α-arabinofuranosidases, cellobiohydrolases, endoglucanases and β-glucosidases, were also identified in the secretome.ConclusionsThis is the first report of a transcriptome and secretome experiment of Aspergillus fumigatus in the degradation of pretreated sugarcane bagasse. The results suggest that this strain employs important strategies for this complex degradation process. It was possible to identify a set of genes and proteins that might be applied in several biotechnology fields. This knowledge can be exploited for the improvement of 2G ethanol production by the rational design of enzymatic cocktails.
Highlights
Sugarcane bagasse has been proposed as a lignocellulosic residue for second-generation ethanol (2G) produced by breaking down biomass into fermentable sugars
To gain more insight into how efficiently A. fumigatus AF293 can depolymerize the sugarcane bagasse, a complex biomass important for Brazilian 2G ethanol production, and to identify genes and proteins responsible for these lignocellulosic breakdown reactions, we examined the transcriptional response by Ribonucleic acid (RNA)-Seq and proteomic profile by mass spectrometry (LC-MS/MS) of A. fumigatus that was cultivated in the presence of steam-exploded sugarcane bagasse (SEB)
Enzymatic analysis To evaluate the activity of enzymes produced by A. fumigatus in the presence of sugarcane bagasse, we performed enzymatic assays using xyloglucan, β-glucan, and CMC as substrates
Summary
Sugarcane bagasse has been proposed as a lignocellulosic residue for second-generation ethanol (2G) produced by breaking down biomass into fermentable sugars. Large amounts of sugarcane straw and bagasse are generated every year in Brazil, so this biomass could be used as a substrate to produce 2G bioethanol, which in a few years will compete with 1G ethanol costs [6,7,8,9,10,11]. The main polymeric component of plant biomass, usually contains regions that are highly crystalline. It is a linear polymeric chain of over 10,000 D-glucose residues linked by β-1,4-glycosidic bonds [12,13,14]. Lignocellulose is difficult to degrade, even when enzymes work synergistically [12, 16]
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