Periplasmic dehydroshikimate dehydratase combined with quinate oxidation in Gluconobacter oxydansfor protocatechuate production

Protocatechuate (3,4-dihydroxybenzoate) has antioxidant properties and is a raw material for the production of muconic acid, which is a key compound in the synthesis of polymers such as nylon and polyethylene terephthalate. Gluconobacter oxydansstrain NBRC3244 has a periplasmic system for oxidation of quinate to produce 3-dehydroquinate. Previously, a periplasmic 3-dehydroshikimate production system was constructed by heterologously expressing Gluconacetobacter diazotrophicusdehydroquinate dehydratase in the periplasm of G. oxydansstrain NBRC3244. 3-Dehydroshikimate is converted to protocatechuate by dehydration. In this study, we constructed a G. oxydansstrain that expresses the Acinetobacter baylyi quiCgene, which encodes a dehydroshikimate dehydratase of which the subcellular localization is likely the periplasm. We attempted to produce protocatechuate by co-cultivation of two recombinant G. oxydansstrains—one expressing the periplasmically targeted dehydroquinate dehydratase and the other expressing A. baylyidehydroshikimate dehydratase. The co-cultivation system produced protocatechuate from quinate in a nearly quantitative manner.Graphical AbstractPeriplasmic metabolic engineering in Gluconobactersp. to expand the intrinsic quinate oxidation to protocatechuate production by heterologous expression of two enzymes.