The compound (2S)-eriodictyol is an important flavonoid that can be derived from (2S)-naringenin through flavonoid 3'-hydroxylase (F3'H) catalyzation. F3'H is a cytochrome P450 enzyme that requires a cytochrome P450 reductase (CPR) to function. However, P450s have limited applications in industrial scale biosynthesis, owing to their low activity. Here, an efficient SmF3'H and a matched SmCPR were identified from Silybum marianum. To improve the efficiency of SmF3'H, we established a high-throughput detection method for (2S)-eriodictyol, in which the promoter combination of SmF3'H and SmCPR were optimized in Saccharomyces cerevisiae. The results revealed that SmF3'H/SmCPR should be expressed by using promoters with similar and strong expression levels. Furthermore, directed evolution was applied to further improve the efficiency of SmF3'H/SmCPR. With the optimized promoter and mutated combinations SmF3'H^D285N/SmCPR^I453V, the (2S)-eriodictyol titer was improved to 3.3 g/L, the highest titer in currently available reports. These results indicated that S. cerevisiae is an ideal platform for functional expression of flavonoid related P450 enzymes.

• PMID: 33226782
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Journal Article | Research Support, Non-U.S. Gov't

Authors

Gao S, Xu X, Zeng W, Xu S, Lyv Y, Feng Y, Kai G, Zhou J, Chen J

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