The Structure of 2-Oxo-4-hydroxy-4-carboxy-5-ureidoimidazoline Decarboxylase Provides Insights into the Mechanism of Uric Acid Degradation

Riccardo Percudani,Rodolfo Berni,Ileana Ramazzina,Giuseppe Zanotti,Claudia Folli,Laura Cendron

doi:10.1074/jbc.m701297200

Riccardo Percudani, Rodolfo Berni + Show 4 more

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https://doi.org/10.1074/jbc.m701297200

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Abstract

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.

Highlights

Uric acid, a key compound in the degradation pathway of purines [1], is degraded in many organisms to (S)-allantoin
We describe the 1.8 Aresolution 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
Of other enzymatic activities associated with urate degradation [4, 5] and to the elucidation of the complete urate degradation pathway [6], involving three enzymatic reactions: oxidation of urate, catalyzed by urate oxidase, yields 5-hydroxyisourate (HIU),3 which is acted upon HIU hydrolase to yield 2-oxo4-hydroxy-4-carboxy-5-ureidoimidazoline (OHCU); in turn, OHCU undergoes stereoselective decarboxylation by the action of OHCU decarboxylase to give CO2 and (S)-allantoin (Scheme 1)

Summary

Introduction

A key compound in the degradation pathway of purines [1], is degraded in many organisms to (S)-allantoin. Crystal Structure of OHCU Decarboxylase in complex with two different ligands and on mutational analysis of the enzyme active site, we provide insights into the mechanism and stereoselectivity of the enzymatic reaction.

Results

Conclusion