Genetic engineering study demonstrates direct cellobionate production in Thermothelomyces heterothallica, indicating a viable platform for enzyme-free biomass conversion.
Key Points
Engineer the thermophilic filamentous fungus Thermothelomyces heterothallica to directly convert alkali-pretreated wheat straw into cellobionic acid without supplementing external cellulase enzymes.
Sequentially disrupted 11 target genes involved in cellobiose and cellobionate catabolism—including eight β-glucosidases, one cellobiose/cellobionate phosphorylase, and two putative cellobionate transporters—using an RNP-based CRISPR-Cas9 system to generate strain TH11.
Cultured the engineered TH11 strain on cellobiose and alkali-pretreated wheat straw, optimizing fermentation parameters including pH (50 mM citrate buffer, pH 6.0) and temperature (43 °C).
The engineered TH11 strain achieved direct conversion of wheat straw, producing 64 mM cellobionate within 5 days.
Substrate conversion efficiency reached approximately 88% cellulose conversion, yielding about 96% cellobionate relative to the consumed cellulose.