ABSTRACT
The glycolytic enzymes of trypanosomes are attractive drug targets, since the blood-stream form of Trypanosoma brucei lacks a functional citric acid cycle and is dependent solely on glycolysis for its energy requirements. Glyceraldehyde-3-phosphate dehydrogenases (GAPDH) from the pathogenic trypanosomatids T. brucei, Trypanosoma cruzi and Leishmania mexicana are quite similar to each other, and yet have sufficient structural differences compared to the human enzyme to enable the structure-based design of compounds that selectively inhibit all three trypanosomatid enzymes but not the human homologue. Adenosine analogs with substitutions on N-6 of the adenine ring and on the 2' position of the ribose moiety were designed, synthesized and tested for inhibition. Two crystal structures of L. mexicana glyceraldehyde-3-phosphate dehydrogenase in complex with high-affinity inhibitors that also block parasite growth were solved at a resolution of 2.6 A and 3.0 A. The complexes crystallized in the same crystal form, with one and a half tetramers in the crystallographic asymmetric unit. There is clear electron density for the inhibitor in all six copies of the binding site in each of the two structures. The L. mexicana GAPDH subunit exhibits substantial structural plasticity upon binding the inhibitor. Movements of the protein backbone, in response to inhibitor binding, enlarge a cavity at the binding site to accommodate the inhibitor in a classic example of induced fit. The extensive hydrophobic interactions between the protein and the two substituents on the adenine scaffold of the inhibitor provide a plausible explanation for the high affinity of these inhibitors for trypanosomatid GAPDHs.