Synthetic Protein Scaffolds for Improving R-(-)-Linalool Production in Escherichia coli

R-(-)-Linalool is widely used in the pharmaceutical, agrochemical, and fragrance industries; however, its applications are limited owing to low yield and high cost of production. To improve the production efficiency of R-(-)-linalool in Escherichia coli, three enzymes [E. coli-derived isopentenyl diphosphate isomerase, Abies grandis-derived geranyl diphosphate synthase, and Streptomyces clavuligerus-derived (3R)-linalool synthases] were physically colocalized to synthetic complexes using synthetic protein scaffolds of GTPase-binding domain, Src homology 3, and PSD95/DlgA/Zo-1. R-(-)-Linalool was produced at the highest concentration in the strain IGL114 containing a scaffold ratio of 1:1:4. By further optimizing the inducer, temperature, and glycerol concentration, the production titer of R-(-)-linalool in the shake flask was increased by approximately 10 times compared with that of the scaffold-free control and was 2.78 times the previously reported yield. The production in the fermenter was about 1.5 times the previous highest production. In general, the final strain accumulated 277.8 and 1523.2 mg/L R-(-)-linalool under the conditions of shake-flask and fed-batch fermentation, respectively. This study provides a foundation for the assembly of bacterial intracellular protein scaffolds.