Human UMP synthase is a bifunctional enzyme that catalyzes the penultimate and last steps in the de novo biosynthesis of UMP. In contrast to prokaryotes, UMP synthase from higher eukaryotes combines the orotate phosphoribosyltransferase and the orotidine-5'-monophosphate (OMP) decarboxylase activities on a single polypeptide chain. The decarboxylase activity is unusual in that it represents the fastest rate acceleration of any enzyme studied to date. Although several crystal structures of OMP decarboxylases have been described, the precise decarboxylation mechanism remains elusive. The crystal structure of the OMP decarboxylase domain from human UMP synthase was determined by molecular replacement using data from a highly twinned monoclinic crystal. The space group is P2(1), with unit-cell parameters a = 69.18, b = 61.70, c = 69.17 A, beta = 113.06 degrees. Self-rotation function analysis and various intensity statistics revealed the presence of pseudo-merohedral twinning, but these tests underestimated the true twin fraction of alpha approximately = 0.44. Data analysis, the origin of the twinning and structure determination are discussed.