Bornyl Diphosphate Synthase From Cinnamomum burmanni and Its Application for (+)-Borneol Biosynthesis in Yeast

Abstract

(+)-Borneol is a desirable monoterpenoid with effective anti-inflammatory and analgesic effects that is known as soft gold. (+)-bornyl diphosphate synthase is the key enzyme in the (+)-borneol biosynthesis pathway. Despite several reported (+)-bornyl diphosphate synthase genes, relatively low (+)-borneol production hinders the attempts to synthesize it using microbial fermentation. Here, we identified the highly specific (+)-bornyl diphosphate synthase CbTPS1 from <i>Cinnamomum burmanni</i>. An <i>in vitro</i> assay showed that (+)-borneol was the main product of CbTPS1 (88.70% of the total products), and the <i>K</i> _<i>m</i> value was 5.11 ± 1.70 μM with a <i>k</i> _<i>cat</i> value of 0.01 s^-1. Further, we reconstituted the (+)-borneol biosynthetic pathway in <i>Saccharomyces cerevisiae</i>. After tailored truncation and adding Kozak sequences, the (+)-borneol yield was improved by 96.33-fold to 2.89 mg⋅L^-1 compared with the initial strain in shake flasks. This work is the first reported attempt to produce (+)-borneol by microbial fermentation. It lays a foundation for further pathway reconstruction and metabolic engineering production of this valuable natural monoterpenoid.