W0419

Homologous Enzymes the Catalyze Unrelated Reactions: 3-keto-L-gulonate 6-phosphate Decarboxylase and Orotidine Monophosphate Decarboxylase. Eric Wise^¶, Wen Shan Yew^⊥, John A. Gerlt^⊥, and Ivan Rayment^¶, ^¶Dept. of Biochemistry, Univ. of Wisconsin, Madison, WI 53706, ^⊥Depts. of Biochemistry and Chemistry, Univ. of Illinois, Urbana, IL 61801.

The divergent evolution of new enzyme functions from existing ones has been widely accepted to occur with either a retention of substrate affinity or a retention of mechanistic feature. Recent investigations into the 3-keto-L-gulonate 6-phosphate decarboxylase (KGPDC) encoded by the UlaD gene in E. Coli and orioidine monophosphate decarboxylase (OMPDC) have suggested that KGPDC and OMPDC are divergently related but share neither a common substrate affinity nor any mechanistic feature. While the OMPDC catalyzed reaction proceeds via a metal ion-independent, concerted mechanism, the KGPDC reaction appears to proceed in a stepwise manner and requires a divalent cation for catalysis. We have determined high-resolution structures of KGPDC by X-ray crystallography with bound analogues of the substrate and proposed intermediate, and the reaction product. Like OMPDC, KGPDC adopts an (α/β)₈ barrel fold and assembles as a homo-dimer. Despite only 20% sequence identity, the structures of the KGPDC and OMPDC dimers overlay almost exactly. The remarkable similarity of the KGPDC structure to the OMPDC structure confirms the evolutionary relationship between these two enzymes. Even more remarkable are the nearly identical active sites of KGPDC and OMPDC. The positions of several conserved active site residues are nearly identical in the KGPDC active site and the OMPDC active site. Although both enzymes share several conserved active site residues, each uses them in a different way to catalyze unrelated reactions. Instead of sharing a common substrate affinity or mechanistic feature, KGPDC and OMPDC share a common active site architecture that each enzyme uses to catalyze a reaction unrelated to the other. Together, OMPDC and KGPDC represent the first known example of enzymes related in this way and may serve as a paradigm for a new mechanism of divergent enzyme evolution.