Fig. 3.

Metabolic engineering of the heterologous APP biosynthetic pathway in E. coli. a Endogenous and heterologous APP biosynthetic pathway in E. coli. b Optimization of the expression of the blue and pduX genes via a bicistronic design that included an MBP tag upstreat of the bluE gene and a his tag upstream of the pduX gene. c Confirmation of the enzymatic functions of PduX and BluE in E. coli. d Confirmation of function of CobD from S. typhimurium in E. coli. e Confirmation of the function of CobC from R. capsulatus in E. coli. Note that Calf intestinal alkaline phosphatase can transform APP to AP. Owing to the lack of a commercially available APP standard, we indirectly monitored the APP level by detecting conversion to AP using the endogenous phosphatase of E. coli