Cendron Laura, Berni Rodolfo, Folli Claudia, Ramazzina Ileana, Percudani Riccardo, Zanotti Giuseppe
Department of Chemistry, University of Padua, and Instituto di Chimica Biomolecolare-Consiglio Nazionale delle Ricerche, Section of Padua, Via Marzolo 1, 35131 Padua, Italy; Venetian Institute of Molecular Medicine, Via Orus 2, 35127 Padua, Italy.
Department of Biochemistry and Molecular Biology, University of Parma, Viale delle Scienze 23/A, 43100 Parma, Italy.
J Biol Chem. 2007 Jun 22;282(25):18182-18189. doi: 10.1074/jbc.M701297200. Epub 2007 Apr 11.
The complete degradation of uric acid to (S)-allantoin, as recently elucidated, involves three enzymatic reactions. Inactivation by pseudogenization of the genes of the pathway occurred during hominoid evolution, resulting in a high concentration of urate in the blood and susceptibility to gout. Here, we describe the 1.8A resolution crystal structure of the homodimeric 2-oxo-4-hydroxy-4-carboxy-5-ureidoimidazoline decarboxylase, which catalyzes the last step in the urate degradation pathway, for both ligand-free enzyme and enzyme in complex with the substrate analogs (R)-allantoin and guanine. Each monomer comprises ten alpha-helices, grouped into two domains and assembled in a novel fold. The structure and the mutational analysis of the active site have allowed us to identify some residues that are essential for catalysis, among which His-67 and Glu-87 appear to play a particularly significant role. Glu-87 may facilitate the exit of the carboxylate group because of electrostatic repulsion that destabilizes the ground state of the substrate, whereas His-67 is likely to be involved in a protonation step leading to the stereoselective formation of the (S)-allantoin enantiomer as reaction product. The structural and functional characterization of 2-oxo-4-hydroxy-4-carboxy-5-ureidoimidazoline decarboxylase can provide useful information in view of the potential use of this enzyme in the enzymatic therapy of gout.
最近的研究表明，尿酸完全降解为（S）-尿囊素涉及三个酶促反应。在类人猿进化过程中，该途径的基因因假基因化而失活，导致血液中尿酸浓度升高，易患痛风。在此，我们描述了同二聚体2-氧代-4-羟基-4-羧基-5-脲基咪唑啉脱羧酶的1.8埃分辨率晶体结构，该酶催化尿酸降解途径的最后一步，包括无配体酶以及与底物类似物（R）-尿囊素和鸟嘌呤结合的酶。每个单体由十个α螺旋组成，分为两个结构域，并以一种新颖的折叠方式组装。活性位点的结构和突变分析使我们能够确定一些对催化至关重要的残基，其中His-67和Glu-87似乎起着特别重要的作用。由于静电排斥作用使底物基态不稳定，Glu-87可能促进羧基的离去，而His-67可能参与导致反应产物（S）-尿囊素对映体立体选择性形成的质子化步骤。鉴于该酶在痛风酶疗法中的潜在应用，对2-氧代-4-羟基-4-羧基-5-脲基咪唑啉脱羧酶的结构和功能表征可以提供有用的信息。
