Structures of human dihydroorotate dehydrogenase in complex with antiproliferative agents

Background: Dihydroorotate dehydrogenase (DHODH) catalyzes the fourth commi tted step in the de novo biosynthesis of pyrimidines. As rapidly proliferat ing human T cells have an exceptional requirement for de novo pyrimidine bi osynthesis, small molecule DHODH inhibitors constitute an attractive therap eutic approach to autoimmune diseases, immunosuppression, and cancer. Neith er the structure of human DHODH nor any member of its family was known. Results: The high-resolution crystal structures of human DHODH in complex w ith two different inhibitors have been solved. The initial set of phases wa s obtained using multiwavelength anomalous diffraction phasing with selenom ethionine-containing DHODH. The structures have been refined to crystallogr aphic R factors of 16.8% and 16.2% at resolutions of 1.6 Angstrom and 1.8 A ngstrom for inhibitors related to brequinar and leflunomide, respectively. Conclusions: Human DHODH has two domains: an alpha/beta-barrel domain conta ining the active site and an alpha-helical domain that forms the opening of a tunnel leading to the active site. Both inhibitors share a common bindin g site in this tunnel, and differences in the binding region govern drug se nsitivity or resistance. The active site of human DHODH is generally simila r to that of the previously reported bacterial active site. The greatest di fferences are that the catalytic base removing the proton from dihydroorota te is a serine rather than a cysteine, and that packing of the flavin monon ucleotide in its binding site is tighter.