Box 16.2 (continued)
dealkylated s-triazine compounds CEAT and CIAT
but not atrazine and of TrzA, a s-triazine hydrolase
(Cook and Hutter 1984), on the other hand, of
Rhodococcus sp. strain TE1 that can degrade atrazine
efficiently to produce the dealkylated metabolites
deisopropylatrazine (CEAT and CIAT) due to AtrA,
an enzyme able to catalyze N-dealkylation reactions.
Based on these reports, it was suggested that in soil,
atrazine was dealkylated under the action of AtrA and
then dealkylated metabolites were dechlorinated by
TrzA. It was recently shown as well that TrzA was
able to transform melamine to ammeline like TriA
(Dodge et al. 2012). It is noteworthy that cloning and
expression of trzA in Rhodococcus sp. strain TE1
successfully led to the dechlorination of the
dealkylated metabolites of atrazine (Shao et al.
1995). The sequence of the gene trzA is known, but
until now the sequence of atrA remained unknown.
This gene was found to be located on a 77 kb plasmid
in Rhodococcus sp. strain TE1 which is able to
degrade atrazine and the thiocarbamate herbicide
EPTC (S-ethyl dipropylcarbamothioate). It was hypothesized that AtrA could be a P450 monooxygenase.
This is also the case for thcB (coding for a P450
monooxygenase), thcC, and thcD genes coding for
EPTC-degrading enzymes in Rhodococcus sp. NI86/
21 which was also shown to degrade atrazine (Nagy
et al. 1995). Similarly, other trz genes coding for
enzymes of s-triazines catabolism have been identified
in Pseudomonas sp. NRRL B-12 227, Pseudomonas
sp. NRRL B-12 228, and Klebsiella pneumoniae.
In the early 1990s, following the observation of
several cases of accelerated degradation of atrazine
in soils repeatedly cropped with corn and consequently
frequently exposed to this herbicide, several bacterial
consortia and even pure bacterial strains able to mineralize this herbicide were isolated (for review see
Udikovic-Kolic et al. 2012). Among these different
atrazine-degrading isolates, Pseudomonas sp. ADP
(Mandelbaum et al. 1995) has been particularly well
studied. In the presence of citrate, this microbial strain
is capable of using atrazine as the sole source of
nitrogen, carbon of atrazine being almost entirely
mineralized. All the genes coding for the enzymes
catalyzing the mineralization of atrazine have been
identified (Martinez et al. 2001). These are the atz
genes coding for a chlorohydrolase (AtzA)
transforming atrazine to hydroxyatrazine, a first
amidohydrolase (AtzB) transforming hydroxyatrazine
(continued)
Box 16.2 (continued)
to N-isopropylammelide and a second one (AtzC) producing cyanuric acid starting from N-isopropylammelide. s-Triazine ring of cyanuric acid is then
degraded by three other enzymes encoded by atzDEF
genes organized as an operon on pADP1 (GarciaGonzalez et al. 2005). Comparison of the conserved
N-terminal sequences rich in histidine residues of
proteins AtzA, B, and C region suggests that these
three genes are derived from a common ancestor. It is
noteworthy that the amino-acid sequence of AtzA
shared only 41 % of similarity with that of TrzA. On
the contrary, the very high similarity (99 %) found
between these genes with atz genes found in various
atrazine-degrading strains suggests their recent divergence. More recently, the trzN gene coding for a
chlorohydrolase has been shown in Nocardioides sp.
C190 (Topp et al. 2000). Although trzN is contributing
to the transformation of atrazine in hydroxyatrazine, it
is significantly differing from atzA and trzA, which
suggest a different origin. This gene has since been
isolated in many bacterial strains and detected in
many agricultural soils cropped with corn and regularly
treated with atrazine (Devers et al. 2007b).
In Pseudomonas sp. ADP, atzABCDEF genes are
located on pADP1, a 96 kb conjugative plasmid,
harboring IncP-1β incompatibility group and most
likely derived from pR751 plasmid. Horizontal gene
transfer (HGT) of pADP1 was studied in vitro, and
thereafter, this phenomenon was observed among
indigenous microflora of a soil cropped with corn
and treated with atrazine (Devers et al. 2005).
In addition, like for phytohormones of synthesis,
HGT of catabolic plasmid harboring atz and trz
genes was suggested following the observation of
these genes on plasmid differing in size detected in
various atrazine-bacterial isolates (Devers et al.
2007a). The observation of IS surrounding atz genes
on these catabolic plasmids further indicates that
genetic rearrangement mediated by IS might occur.
This last hypothesis was otherwise confirmed by the
full sequencing of pADP1 showing that atzA, B, and C
genes are dispersed on the pADP1 plasmid, but each
one is surrounded by IS1071 and IS801 sequences.
The involvement of these IS in the transposition of
atzAB gene cassette by homologous recombination
from pADP1 to the bacterial chromosome was shown
for Variovorax sp. MD1 and MD2 strains (Devers
et al. 2007b). Elsewhere, the atzA gene was already
observed on the chromosome of Arthrobacter sp.
(continued)
696
J.-C. Bertrand et al.
dealkylated s-triazine compounds CEAT and CIAT
but not atrazine and of TrzA, a s-triazine hydrolase
(Cook and Hutter 1984), on the other hand, of
Rhodococcus sp. strain TE1 that can degrade atrazine
efficiently to produce the dealkylated metabolites
deisopropylatrazine (CEAT and CIAT) due to AtrA,
an enzyme able to catalyze N-dealkylation reactions.
Based on these reports, it was suggested that in soil,
atrazine was dealkylated under the action of AtrA and
then dealkylated metabolites were dechlorinated by
TrzA. It was recently shown as well that TrzA was
able to transform melamine to ammeline like TriA
(Dodge et al. 2012). It is noteworthy that cloning and
expression of trzA in Rhodococcus sp. strain TE1
successfully led to the dechlorination of the
dealkylated metabolites of atrazine (Shao et al.
1995). The sequence of the gene trzA is known, but
until now the sequence of atrA remained unknown.
This gene was found to be located on a 77 kb plasmid
in Rhodococcus sp. strain TE1 which is able to
degrade atrazine and the thiocarbamate herbicide
EPTC (S-ethyl dipropylcarbamothioate). It was hypothesized that AtrA could be a P450 monooxygenase.
This is also the case for thcB (coding for a P450
monooxygenase), thcC, and thcD genes coding for
EPTC-degrading enzymes in Rhodococcus sp. NI86/
21 which was also shown to degrade atrazine (Nagy
et al. 1995). Similarly, other trz genes coding for
enzymes of s-triazines catabolism have been identified
in Pseudomonas sp. NRRL B-12 227, Pseudomonas
sp. NRRL B-12 228, and Klebsiella pneumoniae.
In the early 1990s, following the observation of
several cases of accelerated degradation of atrazine
in soils repeatedly cropped with corn and consequently
frequently exposed to this herbicide, several bacterial
consortia and even pure bacterial strains able to mineralize this herbicide were isolated (for review see
Udikovic-Kolic et al. 2012). Among these different
atrazine-degrading isolates, Pseudomonas sp. ADP
(Mandelbaum et al. 1995) has been particularly well
studied. In the presence of citrate, this microbial strain
is capable of using atrazine as the sole source of
nitrogen, carbon of atrazine being almost entirely
mineralized. All the genes coding for the enzymes
catalyzing the mineralization of atrazine have been
identified (Martinez et al. 2001). These are the atz
genes coding for a chlorohydrolase (AtzA)
transforming atrazine to hydroxyatrazine, a first
amidohydrolase (AtzB) transforming hydroxyatrazine
(continued)
Box 16.2 (continued)
to N-isopropylammelide and a second one (AtzC) producing cyanuric acid starting from N-isopropylammelide. s-Triazine ring of cyanuric acid is then
degraded by three other enzymes encoded by atzDEF
genes organized as an operon on pADP1 (GarciaGonzalez et al. 2005). Comparison of the conserved
N-terminal sequences rich in histidine residues of
proteins AtzA, B, and C region suggests that these
three genes are derived from a common ancestor. It is
noteworthy that the amino-acid sequence of AtzA
shared only 41 % of similarity with that of TrzA. On
the contrary, the very high similarity (99 %) found
between these genes with atz genes found in various
atrazine-degrading strains suggests their recent divergence. More recently, the trzN gene coding for a
chlorohydrolase has been shown in Nocardioides sp.
C190 (Topp et al. 2000). Although trzN is contributing
to the transformation of atrazine in hydroxyatrazine, it
is significantly differing from atzA and trzA, which
suggest a different origin. This gene has since been
isolated in many bacterial strains and detected in
many agricultural soils cropped with corn and regularly
treated with atrazine (Devers et al. 2007b).
In Pseudomonas sp. ADP, atzABCDEF genes are
located on pADP1, a 96 kb conjugative plasmid,
harboring IncP-1β incompatibility group and most
likely derived from pR751 plasmid. Horizontal gene
transfer (HGT) of pADP1 was studied in vitro, and
thereafter, this phenomenon was observed among
indigenous microflora of a soil cropped with corn
and treated with atrazine (Devers et al. 2005).
In addition, like for phytohormones of synthesis,
HGT of catabolic plasmid harboring atz and trz
genes was suggested following the observation of
these genes on plasmid differing in size detected in
various atrazine-bacterial isolates (Devers et al.
2007a). The observation of IS surrounding atz genes
on these catabolic plasmids further indicates that
genetic rearrangement mediated by IS might occur.
This last hypothesis was otherwise confirmed by the
full sequencing of pADP1 showing that atzA, B, and C
genes are dispersed on the pADP1 plasmid, but each
one is surrounded by IS1071 and IS801 sequences.
The involvement of these IS in the transposition of
atzAB gene cassette by homologous recombination
from pADP1 to the bacterial chromosome was shown
for Variovorax sp. MD1 and MD2 strains (Devers
et al. 2007b). Elsewhere, the atzA gene was already
observed on the chromosome of Arthrobacter sp.
(continued)
696
J.-C. Bertrand et al.