Active site topology and reaction mechanism of GTP cyclohydrolase I

GTP cyclohydrolase I of Escherichia coli is a torus-shaped homodecamer with D-5 symmetry and catalyzes a complex ring expansion reaction conducive to the formation of dihydroneopterin triphosphate from GTP, The x-ray structure of a complex of the enzyme with the substrate analog, dGTP bound at the active site was determined at a resolution of 3 Angstrom. In the decamer, 10 equivalent active sites are present, each of which contains a 10-Angstrom deep pocket formed by surface areas of 3 adjacent subunits, The substrate forms a complex hydrogen bond network with the protein, Active site residues were modified by site-directed mutagenesis, and enzyme activities of the mutant proteins were measured. On this basis, a mechanism of the enzyme-catalyzed reaction is proposed. Cleavage of the imidazole ring is initiated by protonation of N7 by His-179 followed by the attack of water at CS of the purine system, Cystine Cys-110 Cys-181 may be involved in this reaction step, Opening of the imidazole ring may be in concert with cleavage of the furanose ring to generate a Schiffs base from the glycoside, The gamma-phosphate of GTP may be involved in the subsequent Amadori rearrangement of the carbohydrate side chain by activating the hydroxyl group of Ser-135.