Construction and evolution of anEscherichia colistrain relying on nonoxidative glycolysis for sugar catabolism
The Embden–Meyerhoff–Parnas (EMP) pathway, commonly known as glycolysis, represents the fundamental biochemical infrastructure for sugar catabolism in almost all organisms, as it provides key components for biosynthesis, energy metabolism, and global regulation. EMP-based metabolism synthesizes three-carbon (C3) metabolites before two-carbon (C2) metabolites and must emit one CO2in the synthesis of the C2 building block, acetyl-CoA, a precursor for many industrially important products. Using rational design, genome editing, and evolution, here we replaced the native glycolytic pathways inEscherichia coliwith the previously designed nonoxidative glycolysis (NOG), which bypasses initial C3 formation and directly generates stoichiometric amounts of C2 metabolites. The resulting strain, which contains 11 gene overexpressions, 10 gene deletions by design, and more than 50 genomic mutations (including 3 global regulators) through evolution, grows aerobically in glucose minimal medium but can ferment anaerobically to products with nearly complete carbon conservation. We confirmed that the strain metabolizes glucose through NOG by13C tracer experiments. This redesignedE. colistrain represents a different approach for carbon catabolism and may serve as a useful platform for bioproduction.