Inactivation of triosephosphate isomerase from Trypanosoma cruzi by an agent that perturbs its dimer interface.
We characterized by crystallographic, calorimetric and biochemical methods the action of a low molecular weight compound, 3-(2-benzothiazolylthio)-1-propanesulfonic acid (compound 8) that binds to the dimer interface of triosephosphate isomerase from Trypanosoma cruzi (TcTIM) and thereby abolishes its function with a high level of selectivity. The kinetics of TcTIM inactivation by the agent and isothermal titration calorimetry experiments showed that the binding of two molecules of the compound per enzyme is needed for inactivation. The binding of the first molecule is endothermic, and that of the second exothermic. Crystals of TcTIM in complex with one molecule of the inactivating agent that diffracted to a resolution of 2A were obtained. The compound is at the dimer interface at less than 4A from residues of the two subunits. Compound 8 is more effective at low than at high protein concentrations, indicating that it perturbs the association between the two TcTIM monomers. Calorimetric and kinetic data of experiments in which TcTIM was added to a solution of the inactivating agent showed that at low concentrations of the compound, inactivation is limited by binding, whereas at high concentrations of the agent, the events that follow binding become rate-limiting. The portion of the interface of TcTIM that binds the benzothiazole derivative and its equivalent region in human TIM differs in amino acid composition and hydrophobic packing. Thus, we show that by focusing on protein-protein interfaces, it is possible to discover low molecular weight compounds that are selective for enzymes from parasites.
Instituto de Fisiología Celular, Universidad Nacional Autónoma de México, México, D.F., 04510 México.
Alfredo
A
Téllez-Valencia
Vanesa
V
Olivares-Illana
Alejandra
A
Hernández-Santoyo
Ruy
R
Pérez-Montfort
Miguel
M
Costas
Adela
A
Rodríguez-Romero
Francisco
F
López-Calahorra
Marieta
M
Tuena De Gómez-Puyou
Armando
A
Gómez-Puyou
D017738:Alkanesulfonic Acids / Q000737:chemistry / Q000378:metabolism*; D000818:Animals; D001665:Binding Sites; D002151:Calorimetry; D019281:Dimerization; D006801:Humans; D008958:Models, Molecular; D015394:Molecular Structure; D020836:Protein Structure, Quaternary; D021122:Protein Subunits / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D015800:Protozoan Proteins / Q000737:chemistry* / Q000378:metabolism*; D014305:Triose-Phosphate Isomerase / Q000737:chemistry* / Q000378:metabolism*; D014349:Trypanosoma cruzi / Q000201:enzymology*
D017738:Alkanesulfonic Acids; D021122:Protein Subunits; D015800:Protozoan Proteins; C068559:propylsulfonic acid; D014305:Triose-Phosphate Isomerase