Metabolic engineering of mannitol metabolism to improve protein production in Fusarium venenatum

Abstract

Fusarium venenatum is an important mycoprotein-producing fungus, but the contribution of mannitol metabolism to carbon allocation and fermentation performance remains insufficiently characterized. In this study, single-, double-, and triple-deletion mutants targeting three mannitol metabolism-related genes, the mannitol dehydrogenase gene (FvMDH), the mannitol-1-phosphate dehydrogenase gene (FvM1PDH), and the mannitol metabolism-associated dehydrogenase gene (FvMTDH), were constructed and compared in terms of phenotypes, mannitol accumulation, fermentation performance, and transcriptional responses. FvM1PDH deletion was the main genetic factor associated with reduced mannitol accumulation. The TB6050ΔFvM1PDH strain, hereafter referred to as ΔP, showed 97.60% and 59.62% reductions in cellular and supernatant mannitol, respectively, compared with TB6050. During 5-L bioreactor fermentation, ΔP produced 9.18% more biomass, achieved 10.91% higher glucose conversion efficiency, and increased total crude protein yield per liter by 7.1%, although crude protein content was not significantly changed. Transcriptome analysis revealed changes in central metabolism, including upregulation of the glyoxylate shunt and branched-chain amino acid biosynthesis pathways. Expression of ICL and MS was significantly higher in ΔP than in TB6050 (p_adj < 0.001). In addition, ilvD expression was 2.88-fold higher in ΔP, whereas ACO expression was 2.13-fold lower. Several genes related to oxidative phosphorylation and basal heat response also showed reduced expression. These results support FvM1PDH as a major contributor to mannitol accumulation in F. venenatum and suggest that its deletion is associated with carbon allocation-related metabolic adjustments during glucose-based fermentation.

Keywords

Fusarium venenatum / Mannitol / FvMDH / FvM1PDH / FvMTDH / Transcriptome

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Funding

National Natural Science Foundation of China(32470078)

Central Guidance Fund for Local Science and Technology Development(25ZYCGYS00230)

Tianjin Major Science and Technology Projects and Engineering Programs (Future Industry Projects)

RIGHTS & PERMISSIONS

Jiangnan University