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hydroxyanthranilate 3,4-dioxygenase and 2-amino-3-
carboxymuconate-6-semialdehyde decarboxylase in the
2-nitrobenzoate degradation pathway of Pseudomonas
fluorescens strain KU-7. Appl. Environ. Microbiol., 69,
1564–1572 (2003).
the enzymes and the identification of intermediates
(Tables 1 and 5). In the previously reported pathways
for phenol and its derivatives, catechol is converted to 2-
hydroxymuconic 6-semialdehyde, and then the ring-
cleavage product (2-hydroxymuconic 6-semialdehyde)
is generally metabolized to acetyl-CoA and pyruvate via
a dehydrogenative route and a hydrolytic route.16) Our
results reported here indicate that strain 10d does not
have a 2-hydroxymuconic 6-semialdehyde hydrolase
involved in a hydrolytic route (Table 1). To our knowl-
edge, the phenol-assimilating bacterium Comamonas
teststeroni TA441 metabolizes catechol and the ring-
cleavage product via a meta-cleavage pathway with a
dehydrogenative route only.27) Genetic and biochemical
results indicate that strain TA441 does not have an
NADþ-independent 2-hydroxymuconic 6-semialdehyde
hydrolase. Strain 10d and strain TA441 employ a similar
pathway for the metabolism of 2-hydroxymuconic 6-
semialdehyde.
He and Spain reported that the 2-aminophenol ring-
cleavage pathway is not unique, but is representative of
the metabolic pathways of other 2-aminophenolic com-
pounds.7) As they pointed out, 2-aminophenol and its
methyl-, chloro-, hydroxyl-, and carboxy-derivatives are
metabolized via a 2-aminophenol ring-cleavage pathway
that we designated the modified meta-cleavage pathway.
The metabolic pathway for 4-amino-3-hydroxybenzoic
acid is similar to those for 2-aminophenol derivatives in
the benzene-ring-cleavage step and the deamination
step, but the 4-amino-3-hydroxybenzoic acid metabolic
pathway notably differs from these metabolic pathways
after the deamination step.13) The pathway for 4-amino-
3-hydroxybenozic acid in strain 10d is similar to the
protocatechuic acid pathway in Bacillus circulans in
the oxidative cleavage between positions C2 and C3 of
protocatechuic acid,12,28) but after protocatechuic acid is
converted to 2-hydroxymuconic 6-semialdehyde via 2-
hydroxy-5-carboxymuconic 6-semialdehyde, the 2-hy-
droxymuconic 6-semialdehyde is metabolized via a
dehydrogenative route and a hydrolytic route. We
conclude that the metabolic pathway of 4-amino-3-
hydroxybenzoic acid is different from those of other 2-
aminophenolic compounds.2,3,7)
5) Parales, R. E., Nitrobenzoates and aminobenzoates are
chemoattractants for Pseudomonas strains. Appl. Envi-
ron. Microbiol., 70, 285–292 (2004).
6) Hughes, M. A., and Williams, P. A., Cloning and
characterization of the pnb genes, encoding enzymes
for 4-nitrobenzoate catabolism in Pseudomonas putida
TW3. J. Bacteriol., 183, 1225–1232 (2001).
7) He, Z., and Spain, J. C., Comparison of the downstream
pathways for degradation of nitrobenzene by Pseudo-
monas pseudoalcaligenes JS45 (2-aminophenol path-
way) and by Comamonas sp. JS765 (catechol pathway).
Arch. Microbiol., 171, 309–316 (1999).
8) Davis, J. K., He, Z., Somerville, C. C., and Spain, J. C.,
Genetic and biochemical comparison of 2-aminophenol
1,6-dioxygenase of Pseudomonas pseudoalcaligenes
JS45 to meta-cleavage dioxygenases: divergent evolu-
tion of 2-aminophenol meta-cleavage pathway. Arch.
Microbiol., 172, 330–339 (1999).
9) Takenaka, S., Murakami, S., Kim, Y. J., and Aoki, K.,
Complete nucleotide sequence and functional analysis of
the genes for 2-aminophenol metabolism from Pseudo-
monas sp. AP-3. Arch. Microbiol., 174, 265–272 (2000).
10) Shingler, V., Powlowski, J., and Marklund, U., Nucleo-
tide sequence and functional analysis of the complete
phenol/3,4-dimethylphenol catabolic pathway of Pseu-
domonas sp. strain CF600. J. Bacteriol., 174, 711–724
(1992).
11) Park, H. S., and Kim, H. S., Genetic and structural
organization of the aminophenol catabolic operon and its
implication for evolutionary process. J. Bacteriol., 183,
5074–5081 (2001).
12) Takenaka, S., Asami, T., Orii, C., Murakami, S., and
Aoki, K., A novel meta-cleavage dioxygenase that
cleaves a carboxyl-group-substituted 2-aminophenol:
purification and characterization of 4-amino-3-hydroxy-
benzoate 2,3-dioxygenase from Bordetella sp. strain 10d.
Eur. J. Biochem., 269, 5871–5877 (2002).
13) Orii, C., Takenaka, S., Murakami, S., and Aoki, K., A
novel coupled enzyme assay reveals an enzyme respon-
sible for the deamination of a chemically unstable
intermediate in the metabolic pathway of 4-amino-3-
hydroxybenzoic acid in Bordetella sp. strain 10d. Eur. J.
Biochem., 271, 3248–3254 (2004).
14) Sala-Trepat, J. M., and Evans, W. C., The meta cleavage
of catechol by Azotobacter species: 4-oxalocrotonate
pathway. Eur. J. Biochem., 20, 400–413 (1971).
15) Chen, L. H., Kenyon, G. L., Curtin, F., Harayama, S.,
Bembenek, M. E., Hajipour, G., and Whitman, C. P., 4-
Oxalocrotonate tautomerase, an enzyme composed of 62
amino acid residues per monomer. J. Biol. Chem., 267,
17716–17721 (1992).
16) Harayama, S., Rekik, M., Ngai, K. L., and Ornston, L.
N., Physically associated enzymes produce and metab-
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intermediate formed in catechol metabolism via meta
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