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Maslinic acid conjugate with 7-amino-4-methylcoumarin as probe to monitor the temperature dependent conformational changes of human serum albumin by FRET

2019; Elsevier BV; Volume: 214; Linguagem: Inglês

10.1016/j.saa.2019.02.014

1873-3557

J. A. Molina‐Bolívar, F. Galisteo‐González, C. Carnero Ruiz, Marta Medina‐O’Donnell, Antonio Martı́nez, Andrés Parra,

Lanthanide and Transition Metal Complexes

Synthesis, characterization and spectroscopic investigation of maslinic acid labeled with fluorescent 7-amino-4-methylcoumarin is reported. It was found that the coumarin-maslinic derivative (MaCo) forms an excellent fluorescence resonance energy transfer (FRET) pair with the tryptophan (Trp) residue of human serum albumin (HSA). This feature allowed for monitoring HSA conformational alterations by measuring the distance between donor (Trp) and acceptor (MaCo) through Förster energy transfer mechanism. Displacement experiments confirmed that MaCo binds to subdomain IIA of HSA with independence of temperature. It was observed that, in the temperature range 35-45 °C, the fluorescence emission maximum of HSA-MaCo complex decreased, whereas in the range 45 °C-65 °C, an increment was detected. The concomitant change in the polarity of environment surrounding Trp was confirmed by red edge excitation shift experiments. Thermal denaturation of HSA was followed by time-resolved fluorescence spectroscopy. Average lifetime of Trp residue decreased with temperature due to the increment of solvent collisions and changes in the solvent exposure of Trp. To discriminate the importance of each effect, lifetime of N-Acetyl-L-tryptophanamide (NATA) at different temperatures was measured. Circular dichroism (CD) studies confirmed the loss of secondary structure of HSA with increasing temperature and showed a different trend in the conformational transformation below and above 45 °C, in agreement with steady-state and time-resolved fluorescence experiments.