Selective binding of divalent cations toward heme proteins

Potential toxicity of transition metals like Hg, Cu and Cd are well known and their affinity toward proteins is of great concern. This work explores the selective nature of interactions of Cu2＋, Hg2＋ and Cd2＋ with the heme proteins leghemoglobin, myoglobin and cytochrome C. The binding profiles were analyzed using absorbance spectrum and steady-state fluorescence spectroscopy. Thermodynamic parameters like enthalpy, entropy and free energy changes were derived by isothermal calorimetry and consequent binding parameters were compared for these heme proteins. Free energy （AG） values revealed Cu2＋ binding toward myoglobin and leghemoglobin to be specific and facile in contrast to weak binding for Hg2＋ or Cd2＋ . Time correlated single photon counting indicated significant alteration in excited state lifetimes for metal complexed myoglobin and leghemoglobin suggesting bimolecular collisions to be involved. Interestingly, none of these cations showed significant affinity for cytochrome c pointing that, presence of conserved sequences or heme group is not the only criteria for cation binding toward heme proteins, but the microenvironment of the residues or a specific folding pattern may be responsible for these differential conjugation profile. Binding of these cations may modulate the conformation and functions of these biologically important proteins.

Analysis of curcumin interaction with human serum albumin using spectroscopic studies with molecular simulation

BACKGROUND： Curcumin has emerged to be utilized as a superb beneficial agent, due to its naturally occurring anti-oxidant, anti-inflammatory and anti-carcinogenic property. METHODS： The interaction of curcumin with human serum albumin, the main in vivo transporter of exogenous substances, was investigated using absorption spectroscopy, steady-state fluorescence, excited state life-time studies and circular dichroism spectroscopy.RSULTS：Isothermal titration calorimetry techniques inferred one class of binding site with binding constant -1.74 × 10^5 M ^-1 revealing a strong interaction. The binding profile was analyzed through the evaluation of the thermodynamic parameters, which indicated the involvement of hydrophobic interactions （burial of non-polar group）. Fluorescence lifetime of tryptophan residue was observed to decrease to 1.94 ns from 2.84 ns in presence of Curcumin. Percentage of ct helicity of human serum albumin was also reduced significantly upon binding with curcumin as evidenced by circular dichroism measurement leading to conformational modification of the protein molecule. CONCLUSIONS： On the basis of such complementary results, it may be concluded that curcumin shows strong binding affinity for human serum albumin, probably at the hydrophobic cavities of the protein and at or around the tryptophan residue. Molecular Docking analysis of liSA and curcumin provided light on the number of binding sites at an atomic level, which were already determined at a molecular level in spectroscopic measurements. Our study unfolds the modes of interaction of curcumin with human serum albumin in the light of different biophysical techniques and molecular modeling analysis.