Long-range interactions in the dimer interface of ornithine decarboxylase are important for enzyme function

Ornithine decarboxylase (ODC) is a pyridoxal 5'-phosphate dependent enzyme that catalyzes the first committed step in the biosynthesis of polyamines. ODC is a proven drug target for the treatment of African sleeping sickness. The enzyme is an obligate homodimer, and the two identical active sites ar e formed at the dimer interface. Alanine scanning mutagenesis of dimer inte rface residues in Trypanosoma brucei ODC was undertaken to determine the en ergetic contribution of these residues to subunit association. Twenty-three mutant enzymes were analyzed by analytical ultracentrifugation, and none o f the mutations were found to cause a greater than 1 kcal/mol decrease in d imer stability. These data suggest that the energetics of the interaction m ay be distributed across the interface. Most significantly, many of the mut ations had large effects (Delta DeltaG k(cat)/K-m > 2.5 kcal/mol) on the ca talytic efficiency of the enzyme. Residues that affected activity included those in or near the substrate binding site but also a number of residues t hat are distant (15-20 Angstrom) from this site. These data provide evidenc e that long-range energetic coupling of interface residues to the active si te is essential for enzyme function, even though structural changes upon li gand binding to wild-type ODC are limited to local conformational changes i n the active site. The ODC dimer interface appears to be optimized for cata lytic function and not for dimer stability. Thus, small molecules directed to the ODC interfaces could impact biological function without having to ov ercome the difficult energetic barrier of dissociating the interacting part ners.