Structure of the Cyanuric Acid Hydrolase TrzD Reveals Product Exit Channel

Abstract

Cyanuric acid hydrolases are of industrial importance because of their use in aquatic recreational facilities to remove cyanuric acid, a stabilizer for the chlorine. Degradation of excess cyanuric acid is necessary to maintain chlorine disinfection in the waters. Cyanuric acid hydrolase opens the cyanuric acid ring hydrolytically and subsequent decarboxylation produces carbon dioxide and biuret. In the present study, we report the X-ray structure of TrzD, a cyanuric acid hydrolase from Acidovorax citrulli. The crystal structure at 2.19 Å resolution shows a large displacement of the catalytic lysine (Lys163) in domain 2 away from the active site core, whereas the two other active site lysines from the two other domains are not able to move. The lysine displacement is proposed here to open up a channel for product release. Consistent with that, the structure also showed two molecules of the co-product, carbon dioxide, one in the active site and another trapped in the proposed exit channel. Previous data indicated that the domain 2 lysine residue plays a role in activating an adjacent serine residue carrying out nucleophilic attack, opening the cyanuric acid ring, and the mobile lysine guides products through the exit channel.

The Sodium (Iso)Cyanurates Nax[H3–xC3N3O3]·yH2O (x = 1–3, y = 0, 1): A Key‐Series for Understanding the Crystal Chemistry of Metal (Iso)Cyanurates

“…Besides questions of structural chemistry, a thorough characterization of sodium isocyanurates might also be of considerable interest to environmental chemists,, biochemists,, and toxicologists,, as sodium isocyanurates are not only the most common precursors to, but also the main degradation products of chlorinated isocyanuric acid derivatives used as swimming pool disinfectants, as well as likely degradation products of atrazine and related s‐triazine herbicides. The first description of a member of the Na x [H 3– x C 3 N 3 O 3 ] · y H 2 O series is given by Hofmann with Na 3 [C 3 N 3 O 3 ], including synthesis instructions for phase pure products, common impurities, description of crystal morphologies and chemical analysis.…”

Crystal structures of Moorella thermoacetica cyanuric acid hydrolase reveal conformational flexibility and asymmetry important for catalysis

“…Each RMCAH monomer shows two channels that connect the deeply buried active site to the exterior of the protein. As noted for the equivalent channels observed in the other CAH structures previously determined[8, 9, 30] these channels could serve as paths for the substrate/product to the active site. The two channels show contrasting electrostatic properties (Fig 5A).…”

A procedure for removal of cyanuric acid in swimming pools using a cell-free thermostable cyanuric acid hydrolase

“…The six proteins included three represented in Fig. 1 , M. thermoacetica (WT and C46A) (accession ID: WP_011393610.1) (Shi et al., 2019 ), B. diazoefficiens (formerly Bradyrhizobium japonicum USDA110, accession ID: WP_011090016.1) (Seffernick et al., 2012 ) , Acidovorax citrulli (accession ID: WP_031302833.1) (Bera et al., 2017 ), and two putative CAH enzymes from Pseudolabrys sp. Root1462 (accession ID: WP_056911810.1) and Bradyrhizobium sp.…”