Role of mutation Y6F on the binding properties of Schistosoma japonicum glutathione S-transferase.
The role of the hydroxyl group of tyrosine 6 in the binding of Schistosoma japonicum glutathione S-transferase has been investigated by isothermal titration calorimetry (ITC). A site-specific replacement of this residue with phenylalanine produces the Y6F mutant, which shows negative cooperativity for the binding of reduced glutathione (GSH). Calorimetric measurements indicated that the binding of GSH to Y6F dimer is enthalpically driven over the temperature range investigated. A concomitant net uptake of protons upon binding of GSH to Y6F mutant was detected carrying out calorimetric experiments in various buffer systems with different heats of ionization. The entropy change is favorable at temperatures below 26 degrees C for the first site, being entropically favorable at all temperatures studied for the second site. The enthalpy change of binding is strongly temperature-dependent, arising from a large negative DeltaC(o) (p1)=-3.45+/-0.62kJK(-1)mol(-1) for the first site, whereas a small DeltaC(o) (p2)=-0.33+/-0.05kJK(-1)mol(-1) for the second site was obtained. This large heat capacity change is indicative of conformational changes during the binding of substrate.
Yassin
Zeyad
Z
Dpto. de Química Física, Bioquímica y Q. Inorgánica, Facultad de Ciencias Experimentales, Universidad de Almería, La Cañada de San Urbano, Spain.
Ortiz-Salmerón
Emilia
E
Clemente-Jiménez
M José
MJ
Barón
Carmen
C
García-Fuentes
Luis
L
D019943:Amino Acid Substitution / Q000235:genetics*; D000818:Animals; D001665:Binding Sites; D002151:Calorimetry; D005982:Glutathione Transferase / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D007700:Kinetics; D010649:Phenylalanine / Q000235:genetics*; D011485:Protein Binding; D017510:Protein Folding; D012549:Schistosoma japonicum / Q000201:enzymology*; D013696:Temperature; D013816:Thermodynamics; D014443:Tyrosine / Q000235:genetics*; D014508:Urea / Q000494:pharmacology
D014443:Tyrosine; D010649:Phenylalanine; D014508:Urea; D005982:Glutathione Transferase