Abstract
BackgroundFilamentous fungi are excellent lignocellulose degraders, which they achieve through producing carbohydrate active enzymes (CAZymes). CAZyme production is highly orchestrated and gene expression analysis has greatly expanded understanding of this important biotechnological process. The thermophilic fungus Thermoascus aurantiacus secretes highly active thermostable enzymes that enable saccharifications at higher temperatures; however, the genome-wide measurements of gene expression in response to CAZyme induction are not understood.ResultsA fed-batch system with plant biomass-derived sugars d-xylose, l-arabinose and cellobiose established that these sugars induce CAZyme expression in T. aurantiacus. The C5 sugars induced both cellulases and hemicellulases, while cellobiose specifically induced cellulases. A minimal medium formulation was developed to enable gene expression studies of T. aurantiacus with these inducers. It was found that d-xylose and l-arabinose strongly induced a wide variety of CAZymes, auxiliary activity (AA) enzymes and carbohydrate esterases (CEs), while cellobiose facilitated lower expression of mostly cellulase genes. Furthermore, putative orthologues of different unfolded protein response genes were up-regulated during the C5 sugar feeding together with genes in the C5 sugar assimilation pathways.ConclusionThis work has identified two additional CAZyme inducers for T. aurantiacus, l-arabinose and cellobiose, along with d-xylose. A combination of biochemical assays and RNA-seq measurements established that C5 sugars induce a suite of cellulases and hemicellulases, providing paths to produce broad spectrum thermotolerant enzymatic mixtures.
Highlights
Carbohydrate active enzymes (CAZymes) are vital for the conversion of plant polysaccharides to biofuels and bio-based chemicals [1]
Arabinose and cellobiose induce carbohydrate active enzymes (CAZymes) in T. aurantiacus Previously, fed-batch induction experiments led to the identification of d-xylose as an inducer of CAZymes in T. aurantiacus [11]
We chose two additional lignocellulosederived sugars to test as inducers: l-arabinose, which constitutes ~ 10% of beechwood xylan, and cellobiose, which is a common cellulase inducer and the product of cellobiohydrolase [11]
Summary
Carbohydrate active enzymes (CAZymes) are vital for the conversion of plant polysaccharides to biofuels and bio-based chemicals [1]. The thermophilic fungus Thermoascus aurantiacus is a notable host for thermostable CAZyme production [3]. The enzymes of this fungus were found to be more heat stable and effective at deconstructing lignocellulose than enzymes from other thermophilic fungi and demonstrated the release of sugars from pre-treated biomass at comparable levels to the commercial enzymatic mixture CTec at 50 °C. The thermophilic fungus Thermoascus aurantiacus secretes highly active thermostable enzymes that enable saccharifications at higher temperatures; the genome-wide measurements of gene expression in response to CAZyme induction are not understood
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