STAFF PRIORITY TIME XRAY STRUCT ANALYSIS OF INOSINEMONOPHOSPHATE DEHYDROGENASE

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Inosine monophosphate dehydrogenase (IMPDH) catalyzes the NAD-dependent oxidation of inosine monophosphate (IMP) to xanthine monophosphate (XMP). This is the rate-limiting step in purine biosynthesis. Inhibitors of this enzyme have been shown to have anti-tumor, immunosuppressive, and antiparasitic effects. Inhibition of T. foetus IMPDH in vitro arrests the growth of the organism and this inhibition can be overcome by supplementing the medium with GMP. Native, derivative, and complex data were collected at 7-1 at SSRL. Good thimerosal, PCMBS and tetra-chloro-platinate derivative data sets were obtained and used for MIR phasing. Initial low-resolution SIR and MIR electron density maps indicated a flat, tetrameric molecule, consistent with the expected crystal packing where there is one 503 amino-acid monomer in the asymmetric unit, and the tetramer is generated by the crystallographic four-fold operator. Each monomer forms an a/b barrel resembling very closely the a/b barrel of triose phosphate isomerase (TIM barrel). Subsequent phase combination with the MIR and refined partial model phases has allowed us to fit most of the amino acid backbone and side chains. Heavy-atom difference maps indicate clearly 5 cysteine residues that reacted with either PCMBS or thimerosal, and 3 methionine residues that bound platinum chloride. The active site has been identified by the position of an active-site cysteine and the positions of electron density in difference maps calculated from data collected on ligand and inhibitor-soaked crystals.