Achieving sustainable chemical synthesis and a circular economy will require process innovation to minimize or recover existing waste streams. Valorization of lignin biomass can help advance this goal. The utilization of microbial metabolism to funnel complex and heterogeneous lignin-derived feedstocks to tractable starting points for biochemical upgrading has shown promise. However, this approach necessitates the use of non-model microorganisms, which can be challenging to engineer. We have previously demonstrated that one candidate microbe, Acinetobacter baylyi ADP1, is highly engineerable and capable of undergoing rapid design-build-test-learn cycles, making it an ideal candidate for these applications. Here, we utilize these genetic traits and ADP1’s native β-ketoadipate metabolism to convert mock alkali pretreated liquor lignin (APL) to two valuable natural products, vanillin-glucoside and resveratrol. En route, we create strains with up to 22 genetic modifications, including up to 8 heterologously expressed enzymes, all in a scarless and marker-less manner. Our approach leverages existing aromatic species in APL (vanillate, ferulate, and p-coumarate) to create shortened biochemical routes to end products. This work demonstrates ADP1’s potential as a platform for upgrading lignin waste streams and highlights the potential for biosynthetic methods to maximize the existing chemical potential of lignin aromatic monomers.
