Integrative Biosynthetic Gene Cluster Mining to Optimize a Metabolic Pathway to Efficiently Produce l-Homophenylalanine in Escherichia coli.

Mining biosynthetic genes for the exploration of hybrid metabolic pathways is a promising approach in heterologous production of natural and unnatural products. Here, we developed an integrative biosynthetic gene cluster (BGC) mining strategy to engineer the biosynthesis of l-homophenylalanine (l-Hph), an important intermediate for the synthesis of angiotensin-converting enzyme inhibitors. We assembled the putative l-Hph BGCs and integrated phylogenetic analysis with target metabolite abundance mapping to prioritize candidate BGCs. To obtain an effective l-Hph pathway, various combinations of candidate genes from different species were screened in an iterative design-build-test stepwise manner. After the pathway was strength balanced and the metabolic flux was enhanced, engineered Escherichia coli produced 1.41 g/L of l-Hph from glucose in feeding shake-flask fermentation. Our cluster mining strategy enabled optimization of the target metabolic pathway, and it would be promising for production of other valuable products in the postgenomic era.