Biosynthesis of Bacteriochlorophylls and Bacteriochlorophyllides in Escherichia coli

Photosynthesis, the most important biological process on Earth, converts light energy into chemical energy with essential pigments like chlorophylls and bacteriochlorophylls. The ability to reconstruct photosynthesis in heterotrophic organisms could significantly impact solar energy utilization and biomass production. In this study, we focused on constructing light-dependent biosynthesis pathways for bacteriochlorophyll (BChl) a and bacteriochlorophyllide (BChlide) d and c in the model strain Escherichia coli. The production of the starting compound, Mg protoporphyrin monomethylester, was optimized by screening the ribosome binding sites for the expression of each of the five genes. By fusing a maltose-binding protein and apolipoprotein A-I domain with the membrane protein BchF, the yield of 3-hydroxyethyl-Chlide a was increased by five-fold. Anaerobic cultivation of the engineered E. coli strains facilitated the reduction of the C7=C8 double bond by chlorophyllide a oxidoreductase, a critical step in BChl a synthesis. We further enhanced BChl a production by adjusting the isopropyl-β-d-thiogalactopyranoside concentration to optimize enzyme production and introducing an exogenous superoxide dismutase to combat oxidative stress. Additionally, fusing BciC with a RIAD tag resulted in an eight-fold increase in the production of 3-vinyl BChlide d. This study lays the foundation for the reconstitution of BChl-based photosynthetic apparatus in heterotrophic model organisms, offering promising avenues for future research and applications in biotechnology.