Abstract
Cyanuric acid is a metabolic intermediate of s-triazines, such as atrazine (a common herbicide) and melamine (used in resins and plastics). Cyanuric acid is mineralized to ammonia and carbon dioxide by the soil bacterium Pseudomonas sp. strain ADP via three hydrolytic enzymes (AtzD, AtzE, and AtzF). Here, we report the purification and biochemical and structural characterization of AtzE. Contrary to previous reports, we found that AtzE is not a biuret amidohydrolase, but instead it catalyzes the hydrolytic deamination of 1-carboxybiuret. X-ray crystal structures of apo AtzE and AtzE bound with the suicide inhibitor phenyl phosphorodiamidate revealed that the AtzE enzyme complex consists of two independent molecules in the asymmetric unit. We also show that AtzE forms an α2β2 heterotetramer with a previously unidentified 68-amino acid-long protein (AtzG) encoded in the cyanuric acid mineralization operon from Pseudomonas sp. strain ADP. Moreover, we observed that AtzG is essential for the production of soluble, active AtzE and that this obligate interaction is a vestige of their shared evolutionary origin. We propose that AtzEG was likely recruited into the cyanuric acid-mineralizing pathway from an ancestral glutamine transamidosome that required protein-protein interactions to enforce the exclusion of solvent from the transamidation reaction.
Highlights
Cyanuric acid is a metabolic intermediate of s-triazines, such as atrazine and melamine
We propose that AtzEG was likely recruited into the cyanuric acid–mineralizing pathway from an ancestral glutamine transamidosome that required protein–protein interactions to enforce the exclusion of solvent from the transamidation reaction
The expression of the atzDEF operon is induced under low nitrogen conditions in the presence of cyanuric acid [5], and so we cultured Pseudomonas sp. strain ADP on a minimal medium and supplied cyanuric acid as the sole nitrogen source
Summary
Cyanuric acid is a metabolic intermediate of s-triazines, such as atrazine (a common herbicide) and melamine (used in resins and plastics). Cyanuric acid is mineralized to ammonia and carbon dioxide by the soil bacterium Pseudomonas sp. Strain ADP via three hydrolytic enzymes (AtzD, AtzE, and AtzF). One of the most well-studied examples of an evolutionary response by bacteria to the presence of synthetic xenobiotics is that of the s-triazines [2]. This family of compounds includes fertilizers (e.g. atrazine and ametryn), resins and plastics (e.g. melamine), explosives (e.g. Royal Detonation Explosive), and disinfectants (e.g. cyanuric acid). A common “lower pathway” mineralizes cyanuric acid to carbon dioxide and ammonia
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