Binding of the bioactive component daphnetin to human serum albumin demonstrated using tryptophan fluorescence quenching.
Daphnetin (7,8-dihydroxycoumarin), one of the major bioactive components isolated from Daphne koreane Nakai, has been used in traditional Chinese medicine for the treatment of coagulation disorders. It is also a chelator, an antioxidant and a protein kinase inhibitor. In this paper, a combination of intrinsic fluorescence, Fourier transform infrared (FT-IR) spectroscopy and circular dichroic (CD) spectroscopy has been used to characterize the binding between daphnetin and human serum albumin (HSA) under physiological conditions with drug concentrations of 6.7 x 10(-6) - 2.3 x 10(-5) mol x L(-1), and a HSA concentration of 1.5 x 10(-6) mol x L(-1). Changes in the CD spectra and FT-IR spectra were observed upon ligand binding, and the degree of tryptophan fluorescence quenching did change significantly in the complexes. These data have proved the change in protein secondary structure accompanying ligand binding. The change in tryptophan fluorescence intensity was used to determine the binding constants. The thermodynamic parameters, the enthalpy change (DeltaH) and the entropy change (DeltaS) were calculated to be -12.45 kJ x mol(-1)and 52.48 J x mol(-1) x K(-1) according to the van't Hoff equation, which indicated that hydrophobic and electrostatic interactions played the main role in the binding of daphnetin to HSA, in accordance with the results of calculations performed on a Silicon Graphics Ocatane2 workstation. In addition, the binding distance between daphnetin and HSA was obtained (4.02 nm) based on the Forster energy transfer theory.
Liu
Jiaqin
J
Mianyang Teacher's College, Mianyang 621000, PR China.
Tian
Jianniao
J
Li
Ying
Y
Yao
Xiaojun
X
Hu
Zhide
Z
Chen
Xingguo
X
D002942:Circular Dichroism; D005453:Fluorescence; D006801:Humans; D008958:Models, Molecular; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D012709:Serum Albumin / Q000737:chemistry / Q000378:metabolism*; D012984:Software; D017550:Spectroscopy, Fourier Transform Infrared; D013696:Temperature; D014364:Tryptophan / Q000378:metabolism*; D014468:Umbelliferones / Q000737:chemistry / Q000378:metabolism*
D012709:Serum Albumin; D014468:Umbelliferones; D014364:Tryptophan; C039952:daphnetin