CRYSTAL STRUCTURE OF THE GUANINE NUCLEOTIDE DISSOCIATION INHIBITOR (GDI) DETERMINED AT 1.81 Å RESOLUTION. Ke Zeng, Isabelle Schalk, Shih-Kwang Wu, Enrico A. Stura, Jeanne Madison, M. Huang, Tandon Anurag, W. E. Balch and Ian A. Wilson, Departments of Molecular Biology and Cell Biology, The Scripps Research Institute, 10666 N. Torrey Pines Rd., La Jolla, CA 92037

The crystal structure of the bovine a-isoform of guanine nucleotide dissociation inhibitor (GDI) has been determined to a resolution of 1.8Å. GDI functions in the general recycling of Rab proteins that are involved in regulation of membrane vesicular traffic. The structure of GDI consists of two major domains. The large domain (I) is folded like a cylinder composed of four ß-sheets. The topology and three dimensional structure of domain I surprisingly resemble those of mono-oxygenases and oxidases. Although GDI structure has the similar groove found in those enzymes for FAD binding, and even the sequence remnant Gly-x-Gly structurally aligned well with the corresponding sequence Gly-x-Gly-x-x-Gly for nucleotide binding in those enzymes, no bound ligand was observed in this groove of GDI structure. The smaller domain (II) of GDI contains only a- helices and forms a V-shaped structure with domain I. The three dimensional structure of GDI has distinct regions corresponding to the sequence conserved regions (SCRs) that are common to the choroideraemia (CHM) gene product functioning to deliver Rab to catalytic subunits of Rab geranylgenranyltransferase II. The distribution of these conserved regions suggests that the folding of CHM and GDI may be similar. Site-directed mutagenesis of a region defined by two of the SCRs which forms a compact structure at the apex of domain I reveals a critical role for this region in the binding of Rab proteins. The crystal structure of GDI has been determined using MIRAS method and refined to crystallographic R-value of 19.8% and R-free 27.5% for a total of 430 residues of the molecule in the asymmetric unit.