The response of Pseudomonas putida to a complex lignolysate

There is strong interest in the valorization of lignin derived from plant biomass to produce valuable products; however, the structural complexity of this biopolymer has been a major bottleneck to conversion. Chemical pretreatment liberates soluble fractions of lignin that may be upgraded by biological conversion. Here, ionic liquid pretreatment was employed to obtain soluble aromatic-rich fractions from sorghum, which were converted by Pseudomonas putida KT2440, a promising host for bioconversion of aromatics derived from lignin. Growth studies and mutational analysis demonstrated that P. putida growth on these soluble lignin-derived fractions, referred to as lignolysate, was dependent on aromatic monomers derived from lignin (p-coumarate and ferulate), but other, unknown factors in the lignolysate contributed to growth. Proteomic and metabolomic analyses provided evidence that these unknown factors were amino acids and residual ionic liquid. Proteomic measurements indicated a coordinated response in which these substrates were catabolized simultaneously. A cholinium catabolic pathway was identified and deletion of five genes in the pathway abrogated the ability of P. putida to grow on cholinium as a sole carbon source. This work demonstrates that lignolysates obtained through biomass pretreatment contain multiple substrates and conversion strategies for lignin-derived should take this complexity into account.