Monocyclic Aromatic Hydrocarbon Degradation by Rhodococcus sp. Strain DK17

Abstract

Rhodococcus sp. strain DK17 was isolated from soil and analyzed for the ability to grow on o-xylene as the sole carbon and energy source. Although DK17 cannot grow on m-and p-xylene, it is capable of growth on benzene, phenol, toluene, ethylbenzene, isopropylbenzene, and other alkylbenzene isomers. One UV-generated mutant strain, DK176, simultaneously lost the ability to grow on o-xylene, ethylbenzene, isopropylbenzene, toluene, and benzene, although it could still grow on phenol. The mutant strain was also unable to oxidize indole to indigo following growth in the presence of o-xylene. This observation suggests the loss of an oxygenase that is involved in the initial oxidation of the (alkyl)benzenes tested. Another mutant strain, DK180, isolated for the inability to grow on o-xylene, retained the ability to grow on benzene but was unable to grow on alkylbenzenes due to loss of a meta-cleavage dioxygenase needed for metabolism of methyl-substituted catechols. Further experiments showed that DK180 as well as the wild-type strain DK17 have an ortho-cleavage pathway which is specifically induced by benzene but not by o-xylene. These results indicate that DK17 possesses two different ring-cleavage pathways for the degradation of aromatic compounds, although the initial oxidation reactions may be catalyzed by a common oxygenase. Gas chromatography-mass spectrometry and 300-MHz proton nuclear magnetic resonance spectrometry clearly show that DK180 accumulates 3,4-dimethylcatechol from o-xylene and both 3-and 4-methylcatechol from toluene. This means that there are two initial routes of oxidation of toluene by the strain. Pulsed-field gel electrophoresis analysis demonstrated the presence of two large megaplasmids in the wild-type strain DK17, one of which (pDK2) was lost in the mutant strain DK176. Since several other independently derived mutant strains unable to grow on alkylbenzenes are also missing pDK2, the genes encoding the initial steps in alkylbenzene metabolism (but not phenol metabolism) appear to be present on this approximately 330-kb plasmid.

Benzoate Catabolite Repression of the Phthalate Degradation Pathway in Rhodococcus sp. Strain DK17

“…strain DK17 was originally isolated for the ability to grow on o-xylene and has the capability to utilize such aromatic compounds as benzene, alkylbenzenes (toluene, ethylbenzene, isopropylbenzene, and n-propyl-to n-hexylbenzenes), phenol, and phthalates as sole carbon and energy sources (2,14,15,16,17). This catabolic versatility led us to investigate the adaptability of DK17 to the presence of multiple aromatic hydrocarbons as carbon and energy sources.…”

“…Taken In order to better address the issue of benzoate repression on phthalate utilization by DK17, attempts were made to generate mutant strains defective in the metabolism of benzoate. UV mutagenesis was performed according to the method of Carlton et al (1) with slight modification as described previously (17). After approximately 1,000 colonies were screened, one mutant strain, designated KC710, was isolated for the inability to grow on benzoate as well as the inability to utilize benzoate.…”

Identification of a Novel Dioxygenase Involved in Metabolism ofo-Xylene, Toluene, and Ethylbenzene byRhodococcussp. Strain DK17

“…strain DK17 was originally isolated in Yeochon, Korea, for the ability to grow on o-xylene and was found to also have the capability to grow on benzene, toluene, ethylbenzene, isopropylbenzene, and n-propyl-to n-hexylbenzenes (16). The degradation of o-xylene and toluene in Rhodococcus sp.…”

Functional Identification of OphR, an IclR Family Transcriptional Regulator Involved in the Regulation of the Phthalate Catabolic Operon in Rhodococcus sp. Strain DK17

“…strains DK176 and KC810 were derived from DK17 by curing pDK2 [11] and pDK3 [7], respectively. Initially, DK176 and KC810 were grown overnight in 50 ml mineral salts basal (MSB) medium containing 20 mM glucose.…”

“…As explained in the Introduction, the pDK2-encoded and pDK3-encoded phthalate operons are equally functional in DK17. Accordingly, two previously constructed mutant strains, the pDK2-cured DK176 [11] and the pDK3-cured KC810 [7], were grown on glucose and phthalate, respectively, and the total cellular protein patterns were compared. The soluble cell extracts were separated on onedimensional SDS-PAGE (data not shown).…”