[P001] Poster

12:10 PM - 1:00 PM JST(3:10 AM - 4:00 AM UTC)

[P01-069] Lignin valorization to protocatechuic acid using engineered Acinetobacter baylyi ADP1

○Prashant Singh¹, Guhan Jayaraman¹ (1. Indian Institute of Technology, Madras, Chennai, India (India))

Keywords:

Lignin Valorization,Metabolic Engineering,Protocatechuate,Acinetobacter baylyi ADP1,β-ketoadipate pathway

Lignocellulosic biomass contains lignin, a recalcitrant component that only specific microorganisms can effectively decompose. Acinetobacter baylyi ADP1 is a Gram-negative, naturally competent soil bacterium that metabolizes lignin-related aromatics (LRAs) through the β-ketoadipate pathway. While wild-type ADP1 typically grows on low LRA concentrations (< 20 mM), previous adaptive laboratory evolution (ALE) yielded variants capable of utilizing 100 mM concentrations. Genomic analysis indicated that over 50% of mutations in these evolved strains resulted from insertion sequence (IS) elements.Consequently, we employed an ADP1ΔIS strain as a stable genomic platform for targeted lignin valorization. This study engineered the ADP1ΔIS strain to overproduce protocatechuate (PCA). PCA is a high-value compound used as a polymer precursor, food preservative, and therapeutic agent. Our metabolic strategy employed a dual-substrate approach to drive production. Initial flask-scale experiments using minimal media with coumaric acid and acetate established a baseline PCA yield.To enhance this output, we leveraged the natural competence and homologous recombination capabilities of ADP1 to delete the pcaHG gene, which encodes protocatechuate 3,4-dioxygenase. This genetic modification resulted in a 10-fold increase in PCA yield. Current research focuses on optimizing these yields through integrated metabolic and bioprocess engineering. Future efforts will target additional enzymatic steps within the β-ketoadipate pathway. The objective is to maximize PCA recovery from LRAs and depolymerized lignin fractions derived from corncob.