Mutants of Pseudomonas cepacia G4 defective in catabolism of aromatic compounds and trichloroethylene

Abstract

Pseudomonas cepacia G4 possesses a novel pathway of toluene catabolism that is shown to be responsible for the degradation of trichloroethylene (TCE). This pathway involves conversion of toluene via o-cresol to 3-methylcatechol. In order to determine the enzyme of toluene degradation that is responsible for TCE degradation, chemically induced mutants, blocked in the toluene ortho-monooxygenase (TOM) pathway of G4, were examined. Mutants of the phenotypic class designated TOM Awere all defective in their ability to oxidize toluene, o-cresol, m-cresol, and phenol, suggesting that a single enzyme is responsible for conversion of these compounds to their hydroxylated products (3-methylcatechol from toluene, o-cresol, and m-cresol and catechol from phenol) in the wild type. Mutants of this class did not degrade TCE. Two other mutant classes which were blocked in toluene catabolism, TOM B-, which lacked catechol-2,3-dioxygenase, and TOM C-, which lacked 2-hydroxy-6-oxoheptadienoic acid hydrolase activity, were fully capable of TCE degradation. Therefore, TCE degradation is directly associated with the monooxygenation capability responsible for toluene, cresol, and phenol hydroxylation.

Identifying the genetic basis of ecologically and biotechnologically useful functions of the bacterium Burkholderia vietnamiensis

“…Phenol mutants were identified which showed comparable growth to wild-type G4 on glucose, but reduced or no growth on phenol. Sometimes this phenotype was also associated with a diffuse black pigment likely to be the oxidized product of accumulated catechol (Shields et al, 1991). It should be noted that the mutants tested in the agar screen were distinct from those tested in the STM screen.…”

“…Of the 59 Class Phe TOM mutants, 23 were STM-derived (1.2% hit rate) and 36 were agar-derived (0.6% hit rate). Phe TOM mutations were localized to a 31 172 bp contig of the draft G4 genome sequence which encoded the previously described toluene ortho-monooxygenase (tomA012345; TomA) and catechol 2,3-dioxygenase (tomB; TomB) enzymes (Shields et al, 1991). A putative partition protein (ParA) was encoded upstream of these catabolic genes indicating the contig represented part of the 108 kb catabolic plasmid pTOM (Shields et al, 1995).…”

“…The utility of STM as a method to probe environmental interactions of bacteria was validated by the identification of known pathway components such as tomA/B (Shields et al, 1991) in the phenol-STM screen and rhizosphere competence traits such as amino acid biosynthesis (Rainey, 1999) in the rhizosphere-STM screen. The application of STM also considerably expanded knowledge of the B. vietnamiensis G4 phenol and toluene catabolic pathway, its regulation, and also revealed phenol-specific 'accessory' genes not previously implicated in successful bioremediation.…”

Application of arecAgene-based identification approach to the maize rhizosphere reveals novel diversity inBurkholderiaspecies

“…Many Burkholderia species have been isolated from soil and have been reported to be closely associated with the plant rhizosphere (Parke & Gurian-Sherman, 2001;Coenye & Vandamme, 2003). Beneficial environmental interactions of Burkholderia species include their ability to facilitate both plant protection and growth promotion as biopesticidal agents (Parke & Gurian-Sherman, 2001), the capacity of certain species to fix nitrogen (Minerdi et al, 2001;Caballero-Mellado et al, 2004;Reis et al, 2004), and versatile catabolic capacity that allows degradation of numerous major pollutants such as trichloroethylene and polychlorinated biphenyls (Shields et al, 1991;Goris et al, 2004). In contrast, several Burkholderia species may also cause disease in vulnerable humans , animals and plants (Coenye & Vandamme, 2003).…”

Degradation of mixtures of aromatic and chloroaliphatic hydrocarbons by aromatic hydrocarbon-degrading bacteria

“…The results of degradation experiments depicted in Ta-ble 1 show that G4 and PKO1 were the only strains to completely or nearly completely degrade TCE, BZ, TOL, EBZ, STYR, and all three isomers of xylene, while KR1 degraded a lesser amount of TCE and the xylenes (K = 0.10 for degradation of o-xylene by KR1). These strains, which all express regiospeci¢c monooxygenases, were previously shown to utilize BZ, TOL, and EBZ as sole sources of carbon and energy [7,8,31,32], and to co-oxidize TCE (see Section 1). The oxidation of STYR and all three isomers of xylene by the TOL 3-monooxygenase of PKO1 was also demonstrated previously [8].…”