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[Enzyme Catalysis and Regulation] Identification of Aspartic Acid and Histidine Residues Mediating the Reaction Mechanism and the Substrate Specificity of the Human UDP-glucuronosyltransferases 1A

[Enzyme Catalysis and Regulation] Identification of Aspartic Acid and Histidine Residues Mediating the Reaction Mechanism and the Substrate Specificity of the Human UDP-glucuronosyltransferases 1A - Science News and Views

[Enzyme Catalysis and Regulation] Identification of Aspartic Acid and Histidine Residues Mediating the Reaction Mechanism and the Substrate Specificity of the Human UDP-glucuronosyltransferases 1A - The latest news and publications from Nature, Science, Cell and other journals. Post science topics and your thoughts here. Anything science related goes here.


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Old 12-14-2007, 11:04 AM
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Default [Enzyme Catalysis and Regulation] Identification of Aspartic Acid and Histidine Residues Mediating the Reaction Mechanism and the Substrate Specificity of the Human UDP-glucuronosyltransferases 1A



[Enzyme Catalysis and Regulation] Identification of Aspartic Acid and Histidine Residues Mediating the Reaction Mechanism and the Substrate Specificity of the Human UDP-glucuronosyltransferases 1A

The human UDP-glucuronosyltransferase UGT1A6 is the primary phenol-metabolizing UDP-glucuronosyltransferase isoform. It catalyzes the nucleophilic attack of phenolic xenobiotics on UDP-glucuronic acid, leading to the formation of water-soluble glucuronides. The catalytic mechanism proposed for this reaction is an acid-base mechanism that involves an aspartic/glutamic acid and/or histidine residue. Here, we investigated the role of 14 highly conserved aspartic/glutamic acid residues over the entire sequence of human UGT1A6 by site-directed mutagenesis. We showed that except for aspartic residues Asp-150 and Asp-488, the substitution of carboxylic residues by alanine led to active mutants but with decreased enzyme activity and lower affinity for acceptor and/or donor substrate. Further analysis including mutation of the corresponding residue in other UGT1A isoforms suggests that Asp-150 plays a major catalytic role. In this report we also identified a single active site residue important for glucuronidation of phenols and carboxylic acid substrates by UGT1A enzyme family. Replacing Pro-40 of UGT1A4 by histidine expanded the glucuronidation activity of the enzyme to phenolic and carboxylic compounds, therefore, leading to UGT1A3-type isoform in terms of substrate specificity. Conversely, when His-40 residue of UGT1A3 was replaced with proline, the substrate specificity shifted toward that of UGT1A4 with loss of glucuronidation of phenolic substrates. Furthermore, mutation of His-39 residue of UGT1A1 (His-40 in UGT1A4) to proline led to loss of glucuronidation of phenols but not of estrogens. This study provides a step forward to better understand the glucuronidation mechanism and substrate recognition, which is invaluable for a better prediction of drug metabolism and toxicity in human.
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acid , aspartic , catalysis , enzyme , histidine , human , identification , mechanism , mediating , reaction , regulation , residues , specificity , substrate


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