<i>De novo</i> biosynthesis of phenylpyruvic acid by engineered <i>Escherichia coli</i>

Phenylpyruvic acid is a high-value α-keto acid with broad applications in pharmaceutical and chemical industries. Traditional production via chemical synthesis or transaminase-mediated L-phenylalanine conversion is limited by cost and environmental concerns. Four biosynthetic schemes for phenylpyruvic acid production were evaluated. In scheme a, growing cells converted shikimic acid to phenylpyruvic acid, yielding 0.15 g/L. In scheme b, major transaminases were deleted in an L-phenylalanine producing strain, yielding 0.21 g/L PPA. In scheme c, a "shuttle-run" strategy was applied, with a temperature sensitive pR promoter regulating membrane-associated L-amino acid deaminase, achieving 0.98 g/L. Finally (scheme d), a dual strain "fermentation-enzyme biocatalysis" coupled system combined WP-06-1 with W-03-3, decoupling precursor accumulation from enzymatic conversion and producing 10.21 g/L phenylpyruvic acid, the highest reported from glucose without exogenous L-phenylalanine, providing an efficient and sustainable platform for phenylpyruvic acid biomanufacturing.