| Magnetic Resonance Imaging (MRI) is a clinical technique used for human imaging since 1977. Contrast agents (CA) have been developed that target particular organs and allow for better diagnosis. Gadolinium-based CAs were first developed, followed by iron(III) CAs among others.; To better understand the uptake of these compounds from the blood and their excretion from the body, a study of human glutathione S-transferases (GST), a protein family involved in detoxification, was carried out. Schuhmann-Giampieri proposed that the transhepatic traffic of an hepatobiliary gadolinium(III) CA was determined by the binding capacity of GST. Since some iron(III) CAs are taken up by the liver, similar interactions between iron(III) CAs and GSTs may be anticipated.; Some human GSTs (hGST A1-1, hGST P1-1) have been expressed in E. coli bacteria, purified, characterized, and their non-covalent interactions with some MRI-CAs studied. The experiments included mass spectrometry, enzyme assays, equilibrium dialysis, various spectroscopic measurements and microcalorimetry. Co-crystallization of the hGSTs with the MRI-CAs was also attempted, but unsuccessfully. Mass spectrometry has provided the best evidence for an interaction. A signal corresponding to the mass of one Gd-EOB-DTPA complex "bound" to a protein monomer was detected, as well as a signal for a ratio of up to three Gd(III)-based MRI-CA "bound" per monomer. Thermodynamic parameters were obtained for both Gd-based and Fe-based CAs, using isothermal titration calorimetry and equilibrium dialysis. The interactions were spontaneous reactions, characterized by small negative DeltaG values (DeltaG ≈ -3 kcal/mol). The entropy change (DeltaS) seemed to provide a significant contribution to the interaction. The equilibrium constants revealed weak interactions between the MRI-CAs and the hGSTs, with Kd values in the millimolar range (3.4 mM < Kd < 33 mM).; From this work, it seems that Fe-DESAC and Gd-EOB-DTPA have different behaviors and types of interactions with the human GST enzymes. Whereas the Gd-EOB-DTPA could be transported by the hGSTs in the liver it seems unlikely that an efficient transport of Fe-DESAC would be mediated by the same mechanism. The Fe-DESAC complex seems to have a disruptive effect on the enzyme, as was observed using differential scanning calorimetry, unlike the Gd-EOB-DTPA complex. |
