ادغام مهندسی تشکیل اگزوژن و آندوژن در .E. coli
Engineered assimilation of exogenous and endogenous formate in Escherichia coli
Decoupling biorefineries from land use and agriculture is a major challenge. As formate can be produced from various sources, e.g., electrochemical reduction of CO2, microbial formate-assimilation has the potential to become a sustainable feedstock for the bioindustry. However, organisms that naturally grow on formate are limited by either a low biomass yield or by a limited product spectrum. The engineering of a model biotechnological microbe for growth on formate via synthetic pathways represents a promising approach to tackle this challenge. Here, we achieve a critical milestone for two such synthetic formate-assimilation pathways in Escherichia coli. Our engineering strategy involves the division of the pathways into metabolic modules; the activity of each module – providing at least one essential building block – is selected for in a designated auxotrophic strain. We demonstrate that formate can serve as a sole source of all cellular C1-compounds, including the beta-carbon of serine. We further show that by overexpressing the native threonine cleavage enzymes, the entire cellular glycine requirement can be provided by threonine biosynthesis and degradation. Together, we confirm the simultaneous activity of all pathway segments of the synthetic serine‒threonine cycle. We go beyond the formate bio-economy concept by showing that, under anaerobic conditions, formate produced endogenously by pyruvate formate-lyase can replace exogenous formate. The resulting prototrophic strain constitutes a substantial re-wiring of central metabolism in which C1, glycine, and serine metabolism proceed via a unique set of pathways. This strain can serve as a platform for future metabolic-engineering efforts and could further pave the way for investigating the plasticity of metabolic networks.
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