AtzA is the atrazine chlorohydrolase of Pseudomonas sp. ADP, a cytoplasmic homohexameric Fe(2+)-dependent member of the metallo-dependent hydrolase superfamily (ATZ/TRZ family, amidohydrolase fold). It catalyses the first and committed step of atrazine mineralization, hydrolytic displacement of the ring chlorine by water to give hydroxyatrazine plus chloride; labelling in H2(18)O places the oxygen in the product, establishing hydrolytic dechlorination rather than oxygenation. Specificity is narrow: atrazine, simazine and desethylatrazine are substrates, while terbutylazine, desethyldesisopropylatrazine and melamine are not. AtzA is 98% identical to melamine deaminase (TriA), differing at nine residues, five of them in the active site, and the two enzymes have essentially non-overlapping activities. It carries out a genuine physiological function in its host: Pseudomonas sp. ADP uses atrazine as a nitrogen source, and atzA is the entry point of that catabolic pathway.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0005737 cytoplasm | IEA GO_REF:0000044 | ACCEPT | Summary: Keyword-mapped cytoplasmic location for a soluble hexameric amidohydrolase. Reason: AtzA has no signal peptide or transmembrane segment and was purified as a soluble holoenzyme from cell extracts. UniProt assigns cytoplasm by inference (ECO:0000305) rather than from a localization experiment, which is appropriate at this confidence level. |
| GO:0016787 hydrolase activity | IEA GO_REF:0000002 | MARK AS OVER ANNOTATED | Summary: InterPro-derived root hydrolase term, redundant with the experimentally supported specific activity. Reason: GO:0016787 is the top of the hydrolase branch and is an ancestor of the GO:0018788 atrazine chlorohydrolase annotations that two experimental references support. It is not wrong, but it adds nothing once the specific activity is annotated. The domain-level inference is the correct basis for an uncharacterized family member, not for a gene with purified-enzyme kinetics. |
| GO:0016810 hydrolase activity, acting on carbon-nitrogen (but not peptide) bonds | IEA GO_REF:0000002 | REMOVE | Summary: Fold-level InterPro transfer to the wrong bond class; AtzA cleaves a carbon-halide bond, not a carbon-nitrogen bond. Reason: This annotation comes from IPR011059 (metal-dependent hydrolase composite domain), a structural signature shared across the amidohydrolase superfamily, whose best-known members are C-N hydrolases such as deaminases. AtzA's demonstrated reaction displaces the ring chlorine with water, and it has no detectable deaminase activity with melamine or ammeline. GO places the activity under GO:0019120 hydrolase activity, acting on halide bonds, in C-halide compounds, a sibling branch of GO:0016810. This is a clean example of a fold-based IEA that predicts the superfamily's dominant chemistry rather than the member's own reaction. Notably, the 98%-identical paralog TriA does belong in GO:0016810, so the term is right for the family and wrong for this gene. Supporting Evidence: PMID:8759853 AtzA catalyzes the dechlorination of atrazine, simazine, and desethylatrazine but is not active with melamine, terbutylazine, or desethyldesisopropylatrazine. PMID:11274097 AtzA was shown to exclusively catalyze dehalogenation of halo-substituted triazine ring compounds and had no activity with melamine and ammeline. |
| GO:0018788 atrazine chlorohydrolase activity | EXP PMID:22768133 Intramolecular epistasis and the evolution of a new enzymati... | ACCEPT | Summary: Kinetically characterized atrazine dechlorination with no detectable melamine deaminase activity. Reason: This study measured AtzA's catalytic efficiency for atrazine (kcat/Km = 14,600 s-1.M-1) and confirmed no measurable deaminase activity, while reconstructing substitution trajectories between AtzA and TriA. The substrate is a synthetic herbicide introduced in the 1950s, but the activity is this enzyme's selected function in a host that grows on atrazine, so it belongs in core_functions. A xenobiotic substrate is not the same thing as a non-physiological substrate. Supporting Evidence: PMID:22768133 AtzA is an efficient atrazine dechlorinase with no measurable deaminase activity |
| GO:0018788 atrazine chlorohydrolase activity | EXP PMID:8759853 Atrazine chlorohydrolase from Pseudomonas sp. strain ADP: ge... | ACCEPT | Summary: Founding characterization: purified AtzA converts atrazine to hydroxyatrazine, shown hydrolytic by (18)O labelling. Reason: AtzA was purified to homogeneity and assayed in H2(18)O, which placed the label in hydroxyatrazine and distinguished a chlorohydrolase from an oxygenase. Substrate range was mapped directly. This is the primary evidence for the GO term and for its placement under C-halide bond hydrolases. Supporting Evidence: PMID:8759853 The purified enzyme in H2(18)O yielded [18O]hydroxyatrazine, indicating that AtzA is a chlorohydrolase and not an oxygenase. |
| GO:0018788 atrazine chlorohydrolase activity | IEA GO_REF:0000120 | ACCEPT | Summary: Rhea/EC-driven IEA that happens to land on exactly the right term; a useful contrast with the GO:0016616 benzil case. Reason: RHEA:11312 is an exactMatch of GO:0018788, so this automatic annotation reproduces the experimentally supported activity rather than flattening it to a grouping class. The reaction is in a real pathway (UPA00008 atrazine degradation) and the enzyme is physiologically dedicated to it, which is what distinguishes this Rhea-derived annotation from one generated off a surrogate assay substrate. |
| GO:0019381 atrazine catabolic process | IEA GO_REF:0000041 | ACCEPT | Summary: UniPathway-mapped process term matching the host's demonstrated growth substrate. Reason: Pseudomonas sp. ADP mineralizes atrazine to CO2 and ammonia via hydroxyatrazine, and atzA catalyses step 1 of 3 to cyanurate. The process annotation is pathway-level rather than inferred from the activity alone, and the host's catabolic phenotype was established before the gene was cloned. Supporting Evidence: PMID:8759853 Pseudomonas sp. strain ADP metabolizes atrazine to carbon dioxide and ammonia via the intermediate hydroxyatrazine. |
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Download this section (compressed HTML)Q: Which of AtzA and TriA is ancestral? The direction of the atrazine dechlorinase/melamine deaminase divergence is still unresolved, and it determines whether atrazine dechlorination is a newly recruited activity or a retained one.
Q: Should GO annotate the nine-residue AtzA/TriA difference as an example of recent functional divergence? The pair is a rare case where a documented substitution path separates two distinct, physiologically real activities, and fold-based IEAs cannot distinguish them.
Q: Does the IPR011059 to GO:0016810 mapping over-predict carbon-nitrogen hydrolase activity across the amidohydrolase superfamily generally, given that the same domain also supports C-halide and phosphotriester chemistry?
Experiment: Measure AtzA catalytic efficiency against a panel of chloro- and amino-triazines alongside TriA under identical conditions, including s-triazines plausibly present in soil before synthetic herbicides, to test whether any natural triazine is a competent substrate.
Hypothesis: AtzA's progenitor acted on a naturally occurring triazine or related heterocycle, and residual activity on that compound should be detectable.
Type: enzyme kinetics
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