Metabolic Engineering of Escherichia coli for Production of α‑Santalene, a Precursor of Sandalwood Oil

α-Santalene belongs to a class of natural compounds with many physiological functions and medical applications. Advances in metabolic engineering enable non-native hosts (e.g., Escherichia coli) to produce α-santalene, the precursor of sandalwood oil. However, imbalances in enzymatic activity often result in a metabolic burden on hosts and repress the synthetic capacity of the desired product. In this work, we manipulated ribosome binding sites (RBSs) to optimize an α-santalene synthetic operon in E. coli, and the best engineered E. coli NA-IS_3D strain could produce α-santalene at a titer of 412 mg·L^–1. Concerning the observation of the inverse correlation between indole synthesis and α-santalene production, this study speculated that indole-associated amino acid metabolism would be competitive to the synthesis of α-santalene rather than indole toxicity itself. The deletion of tnaA could lead to a 1.5-fold increase in α-santalene production to a titer of 599 mg·L^–1 in E. coli tnaA^– NA-IS_3D. Our results suggested that the optimization of RBS sets of the synthetic module and attenuation of the competitive pathway are promising approaches for improving the production of terpenoids including α-santalene.