Exploring engineered vesiculation by Pseudomonas putida KT2440 for natural product biosynthesis

Abstract

Pseudomonas species have become promising cell factories for the production of natural products due to their inherent robustness. Although these bacteria have naturally evolved strategies to cope with different kinds of stress, many biotechnological applications benefit from engineering of optimised chassis strains with specially adapted tolerance traits. Here, we explored the formation of outer membrane vesicles (OMV) of Pseudomonas putida KT2440. We found OMV production to correlate with the recombinant production of a natural compound with versatile beneficial properties, the tripyrrole prodigiosin. Further, several P. putida genes were identified, whose up‐ or down‐regulated expression allowed controlling OMV formation. Finally, genetically triggering vesiculation in production strains of the different alkaloids prodigiosin, violacein, and phenazine‐1‐carboxylic acid, as well as the carotenoid zeaxanthin, resulted in up to three‐fold increased product yields. Consequently, our findings suggest that the construction of robust strains by genetic manipulation of OMV formation might be developed into a useful tool which may contribute to improving limited biotechnological applications.

Biotransformation Of l‐Tryptophan To Produce Arcyriaflavin A With Pseudomonas putida KT2440

“…[31][32][33] It has recently been shown that OMV release by hosts like E. coli and P. putida can be genetically engineered thereby supporting the recombinant production of hydrophobic natural compounds. [34,35] One approach to such engineering is the downregulation or deletion of genes with an essential function in the formation of the cell envelope. [36][37][38] Thus, two strains with knocked-out genes mlaE and pvdQ, respectively, previously identified as suitable targets for engineering of vesiculation (see Figure S6), [35] were tested for arcyriaflavin A production (Figure 2C).…”

“…[34,35] One approach to such engineering is the downregulation or deletion of genes with an essential function in the formation of the cell envelope. [36][37][38] Thus, two strains with knocked-out genes mlaE and pvdQ, respectively, previously identified as suitable targets for engineering of vesiculation (see Figure S6), [35] were tested for arcyriaflavin A production (Figure 2C).…”

“…P. putida KT2440 was transformed with the plasmids by plate mating (for genomic integration of rebODCP) using E. coli S17-1 as donor [21] or by electroporation as previously described. [35,52] Construction of seamless deletion mutants was based on the SacB system using the suicide vector pNTPS138-R6KT. [53] For detailed description see SI-M2 in the SI.…”

Anaerobic glucose uptake in Pseudomonas putidaKT2440 in a bioelectrochemical system

“…aromatics, acids and. alcohols) based on a variety of carbon substrates (Bitzenhofer et al, 2023;Dvořák & de Lorenzo, 2018;Loeschcke & Thies, 2020;Tiso et al, 2022;. However, P. putida relies solely on oxygen as the terminal electron acceptor.…”

Economical Production of Phenazine-1-carboxylic Acid from Glycerol by Pseudomonas chlororaphis Using Cost-Effective Minimal Medium

“…The microorganisms that can synthesize phenazine compounds mainly include Pseudomonas [6], Streptomyces [7], and Methanosarcina [8]. The fermentation time of Streptomyces is much longer than that of Pseudomonas; thus, its fermentation cycle is longer, which is not conducive to industrial production.…”