LAUSR

In this study, Y. lipolytica was engineered to produce crotonic acid via the butanol-forming route. Firstly, the crotonase and 3-hydroxybutyryl-CoA dehydrogenase genes from Clostridium beijerinckii, and the thioesterase gene from Bacteroides thetaiotaomicron were heterologously expressed in Y. lipolytica, the engineered strain LZJ001 accumulated 62.3 ± 4.2 mg/L of crotonic acid. Secondly, the acetyl-CoA acetyltransferase from Saccharomyces cerevisiae was overexpressed, the derived recombinant strain LZJ002 produced 123.5 ± 6.8 mg/L of crotonic acid. Finally, the pyruvate dehydrogenase from Escherichia coli was additionally expressed, giving the fully engineered strain LZJ004 that produced 220.0 ± 8.2 mg/L of crotonic acid in shaking-flask culture, which represents a 3.5-fold increase over LZJ001 strain. The approach described here paves the way for environmentally friendly and large-scale industrial production of crotonic acid.