Identification and characterization of the vanillin dehydrogenase YfmT in Bacillus subtilis 3NA.

With vanillin as one of the most important flavoring agents, many efforts have been made to optimize its biotechnological production from natural abundant substrates. However, its toxicity against the hosts results in rather low yields and product concentrations. Bacillus subtilis as a soil-dwelling bacterium is a possible lignin-derived compound-degrading microorganism.

Analysis of the molecular response of Pseudomonas putida KT2440 to the next-generation biofuel n-butanol.

Unlabelled: To increase the efficiency of biocatalysts a thorough understanding of the molecular response of the biocatalyst to precursors, products and environmental conditions applied in bioconversions is essential. Here we performed a comprehensive proteome and phospholipid analysis to characterize the molecular response of the potential biocatalyst Pseudomonas putida KT2440 to the next-generation biofuel n-butanol. Using complementary quantitative proteomics approaches we were able to identify and quantify 1467 proteins, corresponding to 28% of the total KT2440 proteome.

Genetic engineering of Pseudomonas putida KT2440 for rapid and high-yield production of vanillin from ferulic acid.

Vanillin is one of the most important flavoring agents used today. That is why many efforts have been made on biotechnological production from natural abundant substrates. In this work, the nonpathogenic Pseudomonas putida strain KT2440 was genetically optimized to convert ferulic acid to vanillin.

Functional characterization and application of a tightly regulated MekR/P mekA expression system in Escherichia coli and Pseudomonas putida.

A methyl ethyl ketone (MEK)-inducible system based on the broad-host-range plasmid pBBR1MCS2 and on the P mekA promoter region of the MEK degradation operon of Pseudomonas veronii MEK700 was characterized in Escherichia coli JM109 and Pseudomonas putida KT2440. For validation, β-galactosidase (lacZ) was used as a reporter. The novel system, which is positively regulated by MekR, a member of the AraC/XylS family of regulators, was shown to be subject to carbon catabolite repression by glucose, which, however, could not be attributed to the single action of the global regulators Crc and PtsN.

Ethylene glycol metabolism by Pseudomonas putida.

In this study, we investigated the metabolism of ethylene glycol in the Pseudomonas putida strains KT2440 and JM37 by employing growth and bioconversion experiments, directed mutagenesis, and proteome analysis. We found that strain JM37 grew rapidly with ethylene glycol as a sole source of carbon and energy, while strain KT2440 did not grow within 2 days of incubation under the same conditions. However, bioconversion experiments revealed metabolism of ethylene glycol by both strains, with the temporal accumulation of glycolic acid and glyoxylic acid for strain KT2440.

Development of a method for markerless gene deletion in Pseudomonas putida.

We developed a negative counterselection system for Pseudomonas putida based on uracil phosphoribosyltransferase (UPRTase) and sensitivity against the antimetabolite 5-fluorouracil (5-FU). We constructed a P. putida strain that is resistant to 5-FU and constructed vectors for the deletion of the surface adhesion protein gene, the flagellum biosynthesis operon, and two endonuclease genes.