Functional expression of eukaryotic cytochrome P450s in yeast

• Published in: Biotechnology and bioengineering 2021-03, Vol.118 (3), p.1050-1065
• Main Authors: Jiang, Lihong, Huang, Lei, Cai, Jin, Xu, Zhinan, Lian, Jiazhang
• Format: Article
• Language: English
• Subjects: Animals; Carcinogenesis; Carcinogens; Cytochrome; Cytochrome P-450 Enzyme System - biosynthesis; Cytochrome P-450 Enzyme System - genetics; Cytochrome P450; cytochrome P450 reductase; Cytochromes P450; Electron transfer; Endoplasmic reticulum; eukaryotic cytochrome P450 enzymes; Functionals; Gene Expression; Heme; heterologous expression; Humans; Membrane proteins; Metabolism; Proteins; Recombinant Proteins - biosynthesis; Recombinant Proteins - genetics; Reductases; Saccharomyces cerevisiae - enzymology; Saccharomyces cerevisiae - genetics; Xenobiotics; Yeast; yeast expression systems; Yeasts
• ISSN: 0006-3592; 1097-0290
• DOI: 10.1002/bit.27630

Cytochrome P450 enzymes (P450s) are a superfamily of heme‐thiolate proteins widely existing in various organisms. Due to their key roles in secondary metabolism, degradation of xenobiotics, and carcinogenesis, there is a great demand to heterologously express and obtain a sufficient amount of active eukaryotic P450s. However, most eukaryotic P450s are endoplasmic reticulum‐localized membrane proteins, which is the biggest challenge for functional expression to high levels. Furthermore, the functions of P450s require the cooperation of cytochrome P450 reductases for electron transfer. Great efforts have been devoted to the heterologous expression of eukaryotic P450s, and yeasts, particularly Saccharomyces cerevisiae are frequently considered as the first expression systems to be tested for this challenging purpose. This review discusses the strategies for improving the expression and activity of eukaryotic P450s in yeasts, followed by examples of P450s involved in biosynthetic pathway engineering. Eukaryotic cytochrome P450 enzymes (P450s) play critical roles in multiple cellular metabolic processes, and the functional expression of P450s is vital for mechanistic exploration and biotechnological applications. Here, the authors review different strategies for enhancing the expression and activity of eukaryotic P450s in yeast, that is, codon optimization, N‐terminal modification, protein engineering, coexpression with cytochrome P450 reductase (CPR), and construction of engineered yeast chassis. In addition, the authors discussed future perspectives in genome‐scale engineering, high‐throughput technologies, and data‐driven P450 engineering.