Outer membrane vesicles catabolize lignin-derived aromatic compounds in Pseudomonas putida KT2440

Abstract

Lignin is an abundant and recalcitrant component of plant cell walls. While lignin degradation in nature is typically attributed to fungi, growing evidence suggests that bacteria also catabolize this complex biopolymer. However, the spatiotemporal mechanisms for lignin catabolism remain unclear. Improved understanding of this biological process would aid in our collective knowledge of both carbon cycling and microbial strategies to valorize lignin to value-added compounds. Here, we examine lignin modifications and the exoproteome of three aromatic–catabolic bacteria: Pseudomonas putida KT2440, Rhodoccocus jostii RHA1, and Amycolatopsis sp. ATCC 39116. P. putida cultivation in lignin-rich media is characterized by an abundant exoproteome that is dynamically and selectively packaged into outer membrane vesicles (OMVs). Interestingly, many enzymes known to exhibit activity toward lignin-derived aromatic compounds are enriched in OMVs from early to late stationary phase, corresponding to the shift from bioavailable carbon to oligomeric lignin as a carbon source. In vivo and in vitro experiments demonstrate that enzymes contained in the OMVs are active and catabolize aromatic compounds. Taken together, this work supports OMV-mediated catabolism of lignin-derived aromatic compounds as an extracellular strategy for nutrient acquisition by soil bacteria and suggests that OMVs could potentially be useful tools for synthetic biology and biotechnological applications.

Microbiota functional activity biosensors for characterizing nutrient metabolism in vivo

“…A number of proteome publications have demonstrated that inclusion of this detergent in protein extraction protocols increases peptide identification rates from a wide range of proteins, including peptides found in membrane-associated proteins ( Tanca et al, 2013 ). We have also documented an increase in total peptide identifications, including membrane protein representation, after inclusion of SDS in the extraction method that we have applied to a variety of microbial systems ( Blakeley-Ruiz et al, 2019 ; Nickels et al, 2020 ; Salvachúa et al, 2020 ). While there may still be underrepresentation of some proteins, such as membrane proteins with multiple transmembrane helices, if there is an underrepresentation of proteins encoded by the PULs due to methodology-induced bias, this bias will be inherent and consistent across all samples.…”

Industrial biotechnology of Pseudomonas putida: advances and prospects

“…Therefore, cheap, renewable feedstocks with a high level of impurities, such as glycerol, a by-product from biodiesel production (Poblete-Castro et al 2020b ), and aromatic compounds derived from lignin (Kohlstedt et al 2018 ; Vardon et al 2015 ), can be exploited for production of value-added chemicals. The secretion of outer membrane vesicles (OMVs) has been described as an additional mechanism for extracellular nutrient acquisition (Eberlein et al 2019 ; Salvachúa et al 2020b ). (e) A high tolerance against physicochemical stress, chemical stresses (e.g., heavy metal zinc (Peng et al 2018 ), cadmium (Manara et al 2012 ), arsenic (Cánovas et al 2003 )), solvents (Kusumawardhani et al 2018 ), and oxidative stress has been retraced inter alia to an efficient regulation machinery (Kim and Park 2014 ), secretion systems, trans -isomerization of the cell membrane, and changes in head group composition of cell membrane phospholipids (Ramos et al 2015 ).…”

Emerging Strategies for Modifying Lignin Chemistry to Enhance Biological Lignin Valorization

“…[2b,54,55] A recent study by Salvachúa and co-workers reported that Pseudomonas putida KT2440 secreted a striking amount of outer membrane vesicles (OMVs) in lignin-rich medium. [56] Many enzymes involved in the β-ketoadipate pathway which exhibit activity toward lignin-derived aromatic compounds were enriched in OMVs. The OMVs-mediated catabolism provided a means to access carbon sources that cannot translocate the microbial cell membrane and/or mitigate substrate toxicity by restricting cytoplasmic encounters or altering toxic compounds to less toxic molecules extracellularly, thus finally improving the lignin bioconversion performance.…”

“…[10a,56] These findings gained a deeper understanding of how natural systems utilize complex lignin substrates and could provide guidance on the optimization of enzymemicrobe synergy and the rational design of ligninolytic microbes through synthetic biology. [56,57]…”