Heterologous expression of 1,4-benzoquinone reductases from white-rot fungi in Saccharomyces cerevisiae confers protection against p-benzoquinone toxicity

Abstract Background Ethanolic fermentation by Saccharomyces cerevisiae is a key part of biochemical conversion of lignocellulosic feedstocks. Although abundant, lignocellulose typically requires pretreatment for efficient bioconversion. During pretreatment, inhibitory compounds are formed. p -Benzoquinone ( p -BQ) is a lignin-derived inhibitor that is highly toxic to S. cerevisiae . White-rot fungi, efficient degraders of lignin, produce 1,4-benzoquinone reductases (QRDs), which have been suggested to play a role in defense against oxidative stress caused by quinones formed during lignin biodegradation. The hypothesis that QRDs from white-rot fungi can be used to engineer S. cerevisiae with improved resistance against quinone toxicity was explored in experiments with QRD-encoding genes, with and without an N -terminal putative leader sequence, the function of which has not yet been deciphered. Results Constitutive recombinant expression of QRDs from the white-rot fungi Trametes versicolor and Phanerochaete chrysosporium in S. cerevisiae resulted in increased tolerance against p -BQ toxicity compared to an empty vector control containing insert-less plasmid, with QRD-expressing constructs requiring 34 to 42% less time to reach half-time of maximum growth in p -BQ conditions adverse for the growth of the empty vector control (10 mg L − 1 ). All QRD constructs were able to overcome p -BQ toxicity in concentrations that would otherwise be strongly inhibitory (20 mg L − 1 ). Full-length constructs of both T. versicolor and P. chrysosporium QRD typically exhibited 1.5 times higher specific activity than leaderless counterparts and were better detoxifiers. An inverse correlation between QRD enzymatic activity and the time necessary to reach half-time of maximum growth (R 2 = 0.954) was observed during strongly inhibitory conditions. Transformants expressing QRD exhibited maintained glycerol and ethanol yields with increasing p -BQ concentrations, up to 0.44 g g − 1 ethanol at 10 mg L − 1 p -BQ, whereas the empty vector control exhibited increased glycerol yield and decreased ethanol yield, amounting to 0.15 g g − 1 ethanol at 10 mg L − 1 p -BQ. Conclusions Heterologous expression of QRDs from white-rot fungi in S. cerevisiae confers protection against p -BQ toxicity. This investigation supports a role of QRD in the protection against toxic quinones and proposes a biodetoxification strategy against lignin-derived inhibitory p -BQ which can occur simultaneously with fermentation of sugars into ethanol in S. cerevisiae .