Unraveling the trisubstrate-triproduct reaction mechanisms of wine grape VvCYP76F14 to improve wine bouquet

Bouquet is a captivating wine characteristic that affects consumer consumption. Wine lactone correlates with both precursor ((E)-8-carboxylinalool) concentration and wine age that has the greatest contribution to bouquet. In wine grape, cytochrome P450 VvCYP76F14 can catalyze trisubstrate-triproduct reaction processes (hydroxylation, dehydrogenation and carboxylation) to produce (E)-8-carboxylinalool. However, the exact mechanism of the whole reaction is unclear. Here, we unraveled the multi-catalysis mechanism with the aim of rapid conversion of linalool into (E)-8-carboxylinalool in winemaking. Results showed that the redox partner NADPH cytochrome P450 reductase (VvCPR1) was indispensable for hydroxylation and carboxylation, but not dehydrogenation oxidation. Furthermore, the VvCYP76F14-VvCPR1 complex was introduced in the aging stage of winemaking and results showed that the complex could improve the bouquet by increasing the content of wine lactone and shortening the aging time. Nonetheless, this study reveals the trisubstrate-triproduct reaction mechanism of VvCYP76F14 and the VvCYP76F14/VvCPR1 complex has the potential use for wine bouquet enrichment.