Structure and mechanism of GTP cyclohydrolase I of Escherichia coli.

GTP cyclohydrolases I and I1 catalyze the first committed steps in the pathways of pteridine and flavin biosynthesis [ I ] . A reaction pathway for GTP cyclohydrolase I involving the release of C8 of GTP as formate, followed by Amadori rearrangement and ring closure (see Fig. I), has been proposed earlier [2]. The enzyme was first purified and characterised by Yim and Brown [3] Recombinant GTP cyclohydrolase I of Escherichia coli was crystallised, and the structure was solved by X-ray analysis assisted by freeze etching electron microscopy [4,5]. The protein is a torusshaped decamer with D5 symmetry and with approximate dimensions of 100 A in diameter and 65 A in height. The structure is characterised by a novel 20 strand P-barrel. A pocket at the interface of three adjacent subunits A, A* and B located by X-ray analysis of a dGTP complex was identified as the active site. The topology of amino acid residues at the active site is summarised in Fig 2. The guanine moiety is fixed in a cleft constituted by Ile132A*, Glu152A, GlnlSlA, Hisl79A and His1 12A. His1 12A and Ser135A* serve as hydrogen bridge partners of the desoxyribosyl side chain. The entrance to the pocket is lined by a cluster of five basic amino acid residues, Arg65B, His1 13A, Lys68B. Arg185A and Arg139A*. A highly conserved cystine motif (Cysl81A and Cysl IOA) is in close proximity to C8 of the purine nucleotide substrate.