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Endogenous stress caused by faulty oxidation reactions fosters evolution of 2,4-dinitrotoluene-degrading bacteria.

Pérez-Pantoja D, Nikel PI, Chavarría M, de Lorenzo V - PLoS Genet. (2013)

Bottom Line: DNT mineralizes the xenobiotic compound 2,4-dinitrotoluene (DNT) owing to the catabolic dnt genes borne by plasmid DNT, but the process fails to promote significant growth.Naphthalene, the ancestral substrate of the dioxygenase from which DntA has evolved, also caused significant ROS formation.It is thus plausible that the evolutionary roadmap for biodegradation of xenobiotic compounds like DNT was largely elicited by mutagenic oxidative stress caused by faulty reactions of precursor enzymes with novel but structurally related substrates-to-be.

View Article: PubMed Central - PubMed

Affiliation: Systems and Synthetic Biology Program, Centro Nacional de Biotecnología, CSIC, Campus de Cantoblanco, Madrid, Spain.

ABSTRACT
Environmental strain Burkholderia sp. DNT mineralizes the xenobiotic compound 2,4-dinitrotoluene (DNT) owing to the catabolic dnt genes borne by plasmid DNT, but the process fails to promote significant growth. To investigate this lack of physiological return of such an otherwise complete metabolic route, cells were exposed to DNT under various growth conditions and the endogenous formation of reactive oxygen species (ROS) monitored in single bacteria. These tests revealed the buildup of a strong oxidative stress in the population exposed to DNT. By either curing the DNT plasmid or by overproducing the second activity of the biodegradation route (DntB) we could trace a large share of ROS production to the first reaction of the route, which is executed by the multicomponent dioxygenase encoded by the dntA gene cluster. Naphthalene, the ancestral substrate of the dioxygenase from which DntA has evolved, also caused significant ROS formation. That both the old and the new substrate brought about a considerable cellular stress was indicative of a still-evolving DntA enzyme which is neither optimal any longer for naphthalene nor entirely advantageous yet for growth of the host strain on DNT. We could associate endogenous production of ROS with likely error-prone repair mechanisms of DNA damage, and the ensuing stress-induced mutagenesis in cells exposed to DNT. It is thus plausible that the evolutionary roadmap for biodegradation of xenobiotic compounds like DNT was largely elicited by mutagenic oxidative stress caused by faulty reactions of precursor enzymes with novel but structurally related substrates-to-be.

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DNT mineralization pathway in Burkholderia sp. DNT and organization of the dnt gene cluster.(A) The DNT catabolic route starts with the DntA DNT dioxygenase that hydroxylates the aromatic ring in positions 4 and 5 to yield 4M5NC, releasing, at the same time, the first nitro substituent. The substituted catechol is subsequently mono-oxygenated by the DntB hydroxylase, that eliminates the remaining nitro group in the structure, thereby producing 2H5MQ. The rest of the pathway (executed by DntCDGE) includes a ring cleavage reaction and channeling of the products towards the central metabolism, in which they are finally metabolized. (B) Organization of the dnt gene cluster, including dntR, the regulatory gene, and dntAabcd, encoding the multi-component DntA DNT dioxygenase. Amino acid identity between the orthologous dnt and nag genes is included to illustrate the relatedness of the DntA DNT dioxygenase from Burkholderia sp. DNT and the naphthalene dioxygenase from Ralstonia sp. U2.
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pgen-1003764-g001: DNT mineralization pathway in Burkholderia sp. DNT and organization of the dnt gene cluster.(A) The DNT catabolic route starts with the DntA DNT dioxygenase that hydroxylates the aromatic ring in positions 4 and 5 to yield 4M5NC, releasing, at the same time, the first nitro substituent. The substituted catechol is subsequently mono-oxygenated by the DntB hydroxylase, that eliminates the remaining nitro group in the structure, thereby producing 2H5MQ. The rest of the pathway (executed by DntCDGE) includes a ring cleavage reaction and channeling of the products towards the central metabolism, in which they are finally metabolized. (B) Organization of the dnt gene cluster, including dntR, the regulatory gene, and dntAabcd, encoding the multi-component DntA DNT dioxygenase. Amino acid identity between the orthologous dnt and nag genes is included to illustrate the relatedness of the DntA DNT dioxygenase from Burkholderia sp. DNT and the naphthalene dioxygenase from Ralstonia sp. U2.

Mentions: Environmental bacteria capable of biodegradation of 2,4-dinitrotoluene (DNT) afford an exceptional opportunity to examine the factors at play in the emergence of new metabolic abilities. One of such isolates, Burkholderia sp. strain DNT, was isolated from contaminated surface water of an ammunition waste plant on the basis of utilizing DNT as C and N sources [11]. This capacity is due to the action of a route for catabolism of the nitroaromatic compound (Fig. 1A). The phylogeny of the dnt genes and the biochemical properties of the encoded products indicate that this cluster for DNT biodegradation has originated from a precursor pathway for catabolism of the naturally-occurring hydrocarbon naphthalene (Fig. 1B; [12], [13]). Still, the same data indicates that the pathway is even now evolving since the kinetic coupling of each of the biochemical steps is not well balanced [11], the regulation of the system keeps the same effector profile than the precursor operon [14], there are transposon remnants and vestigial genes in the genetic cluster [12] and the strain hardly grows on the substrate of interest [15]. These are all indicators that this strain has started its itinerary towards DNT catabolism but has not yet found an optimal solution to the multi-tiered problem of its viable degradation.


Endogenous stress caused by faulty oxidation reactions fosters evolution of 2,4-dinitrotoluene-degrading bacteria.

Pérez-Pantoja D, Nikel PI, Chavarría M, de Lorenzo V - PLoS Genet. (2013)

DNT mineralization pathway in Burkholderia sp. DNT and organization of the dnt gene cluster.(A) The DNT catabolic route starts with the DntA DNT dioxygenase that hydroxylates the aromatic ring in positions 4 and 5 to yield 4M5NC, releasing, at the same time, the first nitro substituent. The substituted catechol is subsequently mono-oxygenated by the DntB hydroxylase, that eliminates the remaining nitro group in the structure, thereby producing 2H5MQ. The rest of the pathway (executed by DntCDGE) includes a ring cleavage reaction and channeling of the products towards the central metabolism, in which they are finally metabolized. (B) Organization of the dnt gene cluster, including dntR, the regulatory gene, and dntAabcd, encoding the multi-component DntA DNT dioxygenase. Amino acid identity between the orthologous dnt and nag genes is included to illustrate the relatedness of the DntA DNT dioxygenase from Burkholderia sp. DNT and the naphthalene dioxygenase from Ralstonia sp. U2.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3757077&req=5

pgen-1003764-g001: DNT mineralization pathway in Burkholderia sp. DNT and organization of the dnt gene cluster.(A) The DNT catabolic route starts with the DntA DNT dioxygenase that hydroxylates the aromatic ring in positions 4 and 5 to yield 4M5NC, releasing, at the same time, the first nitro substituent. The substituted catechol is subsequently mono-oxygenated by the DntB hydroxylase, that eliminates the remaining nitro group in the structure, thereby producing 2H5MQ. The rest of the pathway (executed by DntCDGE) includes a ring cleavage reaction and channeling of the products towards the central metabolism, in which they are finally metabolized. (B) Organization of the dnt gene cluster, including dntR, the regulatory gene, and dntAabcd, encoding the multi-component DntA DNT dioxygenase. Amino acid identity between the orthologous dnt and nag genes is included to illustrate the relatedness of the DntA DNT dioxygenase from Burkholderia sp. DNT and the naphthalene dioxygenase from Ralstonia sp. U2.
Mentions: Environmental bacteria capable of biodegradation of 2,4-dinitrotoluene (DNT) afford an exceptional opportunity to examine the factors at play in the emergence of new metabolic abilities. One of such isolates, Burkholderia sp. strain DNT, was isolated from contaminated surface water of an ammunition waste plant on the basis of utilizing DNT as C and N sources [11]. This capacity is due to the action of a route for catabolism of the nitroaromatic compound (Fig. 1A). The phylogeny of the dnt genes and the biochemical properties of the encoded products indicate that this cluster for DNT biodegradation has originated from a precursor pathway for catabolism of the naturally-occurring hydrocarbon naphthalene (Fig. 1B; [12], [13]). Still, the same data indicates that the pathway is even now evolving since the kinetic coupling of each of the biochemical steps is not well balanced [11], the regulation of the system keeps the same effector profile than the precursor operon [14], there are transposon remnants and vestigial genes in the genetic cluster [12] and the strain hardly grows on the substrate of interest [15]. These are all indicators that this strain has started its itinerary towards DNT catabolism but has not yet found an optimal solution to the multi-tiered problem of its viable degradation.

Bottom Line: DNT mineralizes the xenobiotic compound 2,4-dinitrotoluene (DNT) owing to the catabolic dnt genes borne by plasmid DNT, but the process fails to promote significant growth.Naphthalene, the ancestral substrate of the dioxygenase from which DntA has evolved, also caused significant ROS formation.It is thus plausible that the evolutionary roadmap for biodegradation of xenobiotic compounds like DNT was largely elicited by mutagenic oxidative stress caused by faulty reactions of precursor enzymes with novel but structurally related substrates-to-be.

View Article: PubMed Central - PubMed

Affiliation: Systems and Synthetic Biology Program, Centro Nacional de Biotecnología, CSIC, Campus de Cantoblanco, Madrid, Spain.

ABSTRACT
Environmental strain Burkholderia sp. DNT mineralizes the xenobiotic compound 2,4-dinitrotoluene (DNT) owing to the catabolic dnt genes borne by plasmid DNT, but the process fails to promote significant growth. To investigate this lack of physiological return of such an otherwise complete metabolic route, cells were exposed to DNT under various growth conditions and the endogenous formation of reactive oxygen species (ROS) monitored in single bacteria. These tests revealed the buildup of a strong oxidative stress in the population exposed to DNT. By either curing the DNT plasmid or by overproducing the second activity of the biodegradation route (DntB) we could trace a large share of ROS production to the first reaction of the route, which is executed by the multicomponent dioxygenase encoded by the dntA gene cluster. Naphthalene, the ancestral substrate of the dioxygenase from which DntA has evolved, also caused significant ROS formation. That both the old and the new substrate brought about a considerable cellular stress was indicative of a still-evolving DntA enzyme which is neither optimal any longer for naphthalene nor entirely advantageous yet for growth of the host strain on DNT. We could associate endogenous production of ROS with likely error-prone repair mechanisms of DNA damage, and the ensuing stress-induced mutagenesis in cells exposed to DNT. It is thus plausible that the evolutionary roadmap for biodegradation of xenobiotic compounds like DNT was largely elicited by mutagenic oxidative stress caused by faulty reactions of precursor enzymes with novel but structurally related substrates-to-be.

Show MeSH
Related in: MedlinePlus