Enhanced tolerance of Clostridium tyrobutyricum to lignin-derived phenolic acids by overexpressing native reductases

Abstract

Ferulic acid (Fer) and p-coumaric acid (Coum) are major phenolic inhibitors in lignocellulosic hydrolysates that severely hinder the growth and metabolism of Clostridia species. This study demonstrates that the reduction of Fer and Coum to dihydroferulic acid and phloretic acid by Clostridium tyrobutyricum significantly alleviates their toxicity. Overexpression of the dho1 and sdr1 genes, encoding Fer and Coum reductases, respectively, in C. tyrobutyricum can significantly enhance tolerance to these phenolic acids. As a result, the recombinant strain ATCC 25755/ds, which co-overexpresses dho1 and sdr1, exhibited a marked increase in butyrate production compared to the wild-type strain under phenolic acid stress. In fed-batch fermentation with a 1.0 g/L mixture of Fer and Coum (1:1, w/w), ATCC 25755/ds showed a 35.1 % increase in butyrate production and a 61.1 % higher productivity. These results indicate that enhancing phenolic acid reduction can significantly improve Clostridia’s tolerance to phenolic acids, thereby strengthening the biotransformation of lignocellulose hydrolysates.