Structural Evidence for a [4Fe‐5S] Intermediate in the Non‐Redox Desulfuration of Thiouracil

Abstract

We recently discovered a[ Fe-S]-containing protein with in vivo thiouracil desulfidase activity,d ubbed TudS.T he crystal structure of TudS refined at 1.5 resolution is reported; it harbors a[ 4Fe-4S] cluster bound by three cysteines only. Incubation of TudS crystals with 4-thiouracil trapped the cluster with ah ydrosulfide ligand bound to the fourth nonprotein-bonded iron, as established by the sulfur anomalous signal. This indicates that a[ 4Fe-5S] state of the cluster is acatalytic intermediate in the desulfuration reaction. Structural data and site-directed mutagenesis indicate that aw ater molecule is located next to the hydrosulfide ligand and to two catalytically important residues,S er101 and Glu45. This information, together with modeling studies allowu st o propose am echanism for the unprecedented non-redox enzymatic desulfuration of thiouracil, in whicha[4Fe-4S] cluster binds and activates the sulfur atom of the substrate.