Pseudomonas putida

About: Pseudomonas putida is a research topic. Over the lifetime, 6854 publications have been published within this topic receiving 230572 citations.

Degradation of 1,4-dichlorobenzene by enriched and constructed bacteria

Abstract: Three strains, RHO1, R3 and B1, tentatively identified as a Pseudomonas sp., an Alcaligenes sp. and a Pseudomonas sp. which were able to use 1,4-dichlorobenzene as the sole carbon and energy source were isolated from water of the Rhine river and from the sewage plant at Leverkusen-Burrig. A hybrid strain, WR1313, which uses chlorobenzene as the growth substrate, was obtained by mating the benzene-growing Pseudomonas putida strain F1 with strain B13, a Pseudomonas sp. degrading chlorocatechols. Further selection of this strain for growth on 1,4-dichlorobenzene allowed the isolation of strain WR1323. During growth on 1,4-dichlorobenzene the strains released stoichiometric amounts of chloride. The affinity of the organisms to 1,4-dichlorobenzene was measured with strain R3 showing a Ks value of 1.2 mg/l. Respiration data and enzyme activities in cell extracts as well as the isolation of 3,6-dichlorocatechol from the culture fluid are consistent with the degradation of 1,4-dichlorobenzene via 3,6-dichlorocatechol, 2,5-dichloro-cis,cis-muconate, 2-chloro-4-carboxymethylenebut-2-en-4-olide.

The ssu locus plays a key role in organosulfur metabolism in Pseudomonas putida S-313.

Abstract: Pseudomonas putida S-313 can utilize a broad range of aromatic sulfonates as sulfur sources for growth in sulfate-free minimal medium. The sulfonates are cleaved monooxygenolytically to yield the corresponding phenols. miniTn5 mutants of strain S-313 which were no longer able to desulfurize arylsulfonates were isolated and were found to carry transposon insertions in the ssuEADCBF operon, which contained genes for an ATP-binding cassette-type transporter (ssuABC), a two-component reduced flavin mononucleotide-dependent monooxygenase (ssuED) closely related to the Escherichia coli alkanesulfonatase, and a protein related to clostridial molybdopterin-binding proteins (ssuF). These mutants were also deficient in growth with a variety of other organosulfur sources, including aromatic and aliphatic sulfate esters, methionine, and aliphatic sulfonates other than the natural sulfonates taurine and cysteate. This pleiotropic phenotype was complemented by the ssu operon, confirming its key role in organosulfur metabolism in this species. Further complementation analysis revealed that the ssuF gene product was required for growth with all of the tested substrates except methionine and that the oxygenase encoded by ssuD was required for growth with sulfonates or methionine. The flavin reductase SsuE was not required for growth with aliphatic sulfonates or methionine but was needed for growth with arylsulfonates, suggesting that an alternative isozyme exists for the former compounds that is not active in transformation of the latter substrates. Aryl sulfate ester utilization was catalyzed by an arylsulfotransferase, and not by an arylsulfatase as in the related species Pseudomonas aeruginosa.

Role of the crc Gene in Catabolic Repression of the Pseudomonas putida GPo1 Alkane Degradation Pathway

Abstract: Expression of the alkane degradation pathway encoded in the OCT plasmid of Pseudomonas putida GPo1 is induced in the presence of alkanes by the AlkS regulator, and it is down-regulated by catabolic repression. The catabolic repression effect reduces the expression of the two AlkS-activated promoters of the pathway, named PalkB and PalkS2. The P. putida Crc protein participates in catabolic repression of some metabolic pathways for sugars and nitrogenated compounds. Here, we show that Crc has an important role in the catabolic repression exerted on the P. putida GPo1 alkane degradation pathway when cells grow exponentially in a rich medium. Interestingly, Crc plays little or no role on the catabolic repression exerted by some organic acids in a defined medium, which shows that these two types of catabolic repression can be genetically distinguished. Disruption of the crc gene led to a six- to sevenfold increase in the levels of the mRNAs arising from the AlkS-activated PalkB and PalkS2 promoters in cells growing exponentially in rich medium. This was not due to an increase in the half-lives of these mRNAs. Since AlkS activates the expression of its own gene and seems to be present in limiting amounts, the higher mRNA levels observed in the absence of Crc could arise from an increase in either transcription initiation or in the translation efficiency of the alkS mRNA. Both alternatives would lead to increased AlkS levels and hence to elevated expression of PalkB and PalkS2. High expression of alkS from a heterologous promoter eliminated catabolic repression. Our results indicate that catabolic repression in rich medium is directed to down-regulate the levels of the AlkS activator. Crc would thus modulate, directly or indirectly, the levels of AlkS.

Determination of the transcription initiation site and identification of the protein product of the regulatory gene xylR for xyl operons on the TOL plasmid.

Abstract: The xylR gene is a regulatory gene on the TOL plasmid, which acts in a positive manner on xyl operons for degradation of toluene and xylenes in Pseudomonas putida. A DNA fragment containing the xylR promoter region was cloned on promoter-probing vectors, and its nucleotide sequence was determined. The transcription initiation site of the xylR gene was determined in cells of P. putida and Escherichia coli by S1 nuclease and reverse transcriptase mapping. Two initiation sites were detected which were identical in both P. putida and E. coli. The amounts of mRNA synthesized in both bacterial cells were almost the same and independent of the inducers for xyl operons. The consensus sequences for E. coli promoters were found in the region preceding the respective transcription initiation sites. The product of the xylR gene was identified by the maxicell system as a protein with an approximate molecular weight of 67,000.