Pairing of orthogonal chaperones with a cytochrome P450 enhances terpene synthesis in Saccharomyces cerevisiae

The supply of terpenes is often limited by their low extraction yield from natural resources, such as plants. Thus, microbial biosynthesis has emerged as an attractive platform for the production of terpenes. Many strategies have been applied to engineer microbes to improve terpene production capabilities; however, functional expression of heterologous proteins such as cytochrome P450 enzymes (P450s) in microbes is a major obstacle. This study reports the successful pairing of cognate chaperones and P450s for functional heterologous expression in Saccharomyces cerevisiae. This chaperone pairing was exploited to facilitate the functional assembly of the protopanaxadiol (PPD) biosynthesis pathway, which consists of a P450 oxygenase and a P450 reductase redox partner originating from Panax ginseng and Arabidopsis thaliana, respectively. We identified several chaperones required for protein folding in P. ginseng and A. thaliana and evaluated the impact of the coexpression of the corresponding chaperones on the synthesis and activity of PPD biosynthesis enzymes. Expression of a chaperone from P. ginseng (PgCPR5), a cognate of PPD biosynthesis enzymes, significantly increased PPD production by more than 2.5-fold compared with that in the corresponding control strain. Thus, pairing of chaperones with heterologous enzymes provides an effective strategy for the construction of challenging biosynthesis pathways in yeast.