Efficient production of butyric acid from lignocellulosic biomass by revealing the mechanisms of Clostridium tyrobutyricum tolerance to phenolic inhibitors.

Phenolic compounds (PCs) generated during pretreatment of lignocellulosic biomass severely hinder the biorefinery by Clostridia. As a hyperbutyrate-producing strain, Clostridium tyrobutyricum has excellent tolerance to PCs, but its tolerance mechanism is poorly understood. In this study, a comprehensive transcriptome analysis was applied to elucidate the response of C. tyrobutyricum to four typical PCs. The findings revealed that the expression levels of genes associated with PC reduction, HSPs, and membrane transport were significantly altered under PC stress. Due to PCs being reduced to low-toxicity alcohols/acids by C. tyrobutyricum, enhancing the reduction of PCs by overexpressing reductase genes could enhance the strain's tolerance to PCs. Under 1.0 g/L p-coumaric acid stress, compared with the wild-type strain, ATCC 25755/sdr1 exhibited a 31.2 % increase in butyrate production and a 38.5 % increase in productivity. These insights contribute to the construction of PC-tolerant Clostridia, which holds promise for improving biofuel and chemical production from lignocellulosic biomass.

Transcriptome analysis reveals reasons for the low tolerance of Clostridium tyrobutyricum to furan derivatives.

Lignocellulosic biomass is considered the most abundant and renewable feedstock for biobased butyric acid production. However, the furan derivatives (FAs, mainly furfural and 5-hydroxymethylfurfural) generated from the pretreatment of lignocellulose severely inhibit the growth of Clostridium tyrobutyricum, which is the best strain for producing butyric acid. The tolerance mechanism of C. tyrobutyricum to FAs has not been investigated thus far. Here, the response of C. tyrobutyricum ATCC 25755 to FA challenge was first evaluated by using comprehensive transcriptional analysis. The results indicated that the genes related to membrane transport, heat shock proteins, and transcriptional regulation were upregulated under FA stress. However, the expression of almost all genes encoding reductases was not changed, and only the ad gene CTK_RS02625 and the bud gene CTK_RS07810 showed a significant increase of ~ 1.05-fold. Then, the enzyme activity assays indicated that BUD could catalyze the reduction of FAs with relatively low activity and that AD could not participate in the conversion of FAs, indicating that the inability to rapidly convert FAs to their low-toxicity alcohols may be the main reason for the low FA tolerance of C. tyrobutyricum. This research provides insights into the development of FA-tolerant strains, thereby enhancing the bioconversion of lignocellulosic biomass to butyric acid. KEY POINTS: • The response of C. tyrobutyricum to FAs was evaluated for the first time. • Genes encoding membrane transporters and heat shock proteins were triggered by FAs. • A lack of effective FA reductases leads to low FA tolerance in C. tyrobutyricum.