Physiological responses of Pseudomonas putida to formaldehyde during detoxification

Abstract

SummaryPseudomonas putida KT2440 exhibits two formaldehyde dehydrogenases and two formate dehydrogenase complexes that allow the strain to stoichiometrically convert formaldehyde into CO2. The strain tolerated up to 1.5 mM formaldehyde and died in the presence of 10 mM. In the presence of 0.5 mM formaldehyde, a sublethal concentration of this chemical, the growth rate decreased by about 40% with respect to growth in the absence of the toxicant. Transcriptomic analysis revealed that in response to low formaldehyde concentrations, a limited number of genes (52) were upregulated. Based on the function of these genes it seems that sublethal concentrations of HCOH trigger responses to overcome DNA and protein damage, extrude this toxic compound, and detoxify it by converting the chemical to CO2. In strains bearing mutations of the upregulated genes we analysed growth inhibition by 1.5 mM HCOH and killing rates by 10 mM HCOH. Mutants in the MexEF/OprN efflux pump and in the DNA repair genes recA and uvrB were hypersensitive to 10 mM HCOH, the killing rate being three to four orders of magnitude higher than those in the wild‐type strain. Mutants in other upregulated genes died at slightly higher or at similar rates to the parental strain. Regarding growth inhibition, we found that mutants in glutathione biosynthesis, stress response mediated by 2‐hydroxy acid dehydrogenases and two efflux pumps of the MSF family were unable to grow in the presence of 1.5 mM HCOH. In an independent screening test we searched for mutants which were hypersensitive to formaldehyde, but whose expression did not change in response to this chemical. Two mutants with insertions in recD and fhdA were found which were unable to grow in the presence of 1.5 mM HCOH. The recD mutant was hypersensitive to 10 mM HCOH and died at a higher rate than the parental strain.

Redundancy of Enzymes for Formaldehyde Detoxification in Pseudomonas putida

“…1). Early microarray data indicated that the level of expression of the four transcriptional units was independent of the presence of formaldehyde or formate in the culture medium (29). To investigate this further, we first determined the transcription start point of all three operons and PP0328, the monocistronic unit, and then constructed the appropriate fusions to ЈlacZ in the low-copy-number pMP220 plasmid.…”

“…These complexes are glutathione-independent enzymes (24). In a previous study we exposed P. putida KT2440 to sublethal concentrations of HCOH and analyzed global cell responses using microarrays (29). This analysis revealed that expression of the genes potentially able to encode formaldehyde dehydrogenases and formate dehydrogenases did not vary significantly in the presence of the aldehyde, although a number of functions related to the defense against this toxic chemical were induced, including DNA repair enzymes and chaperones that refold damaged proteins (29).…”

“…In a previous study we exposed P. putida KT2440 to sublethal concentrations of HCOH and analyzed global cell responses using microarrays (29). This analysis revealed that expression of the genes potentially able to encode formaldehyde dehydrogenases and formate dehydrogenases did not vary significantly in the presence of the aldehyde, although a number of functions related to the defense against this toxic chemical were induced, including DNA repair enzymes and chaperones that refold damaged proteins (29). It was then argued that formaldehyde dehydro-genases and formate dehydrogenases could be expressed constitutively to safeguard cells against potential damage caused by the internal production of HCOH, a dead-end product in the metabolism of histidine and methoxylated aromatic compounds such as vanillate (2,13,17,30).…”

Genome‐wide identification of tolerance mechanisms toward p‐coumaric acid in Pseudomonas putida

“…Some of the changes that can occur in the membrane, when bacteria are exposed to hydrophobic solvents, include vesicle formation, alteration of phospholipid composition, and reduced permeability of the cell membrane (Heipieper & De Bont, 1994; Nicolaou, Gaida, & Papoutsakis, 2010; Ramos et al, 2002). The mechanisms behind tolerance in the model strain P. putida KT2440 has also been studied for different compounds (Benndorf, Thiersch, Loffhagen, Kunath, & Harms, 2006; Domínguez‐Cuevas, González‐Pastor, Marqués, Ramos, & de Lorenzo, 2006; Fernandez, Conde et al, 2012; Fernandez, Niqui‐Arroyo, Conde, Ramos, & Duque, 2012; Roca, Rodríguez‐Herva, Duque, & Ramos, 2008; Santos, Benndorf, & Sá‐Correia, 2004). …”

Structural and Molecular Genetic Analyses of the Bacterial Carbazole Degradation System

“…On the other hand, the mexEFoprN operon, encoding an efflux pump of the resistance-nodulation-cell division family, [105][106][107][108] was specifically upregulated by pCAR1 carriage in P. putida KT2440 (Fig. 12A), whereas the expression of orthologous operons in other species remained unaltered.…”