Ma Ji, Liu Yuan, Chen Luan, Xie Yue, Wang Li-Yun, Xie Meng-Xia
Analytical & Testing Center of Beijing Normal University, Beijing 100875, PR China.
Analytical & Testing Center of Beijing Normal University, Beijing 100875, PR China.
Food Chem. 2012 May 1;132(1):663-70. doi: 10.1016/j.foodchem.2011.11.023. Epub 2011 Nov 11.
The interaction mechanism of 3,7-dihydroxyflavone (3,7-diHF) and human serum albumin (HSA) was investigated by fluorescence quenching, fluorescence enhancement, steady-state and time-resolved fluorescence emission and UV-vis absorption spectrometry. The binding site of 3,7-diHF on protein was determined by investigating the spectroscopic properties of 3,7-diHF-HSA complex at pH 7.4 and pH 3.5 individually, and confirmed by the site marker competitive experiments. The binding parameters of 3,7-diHF-HSA complex were estimated by fluorescence quenching experiments, and the data were in good agreement with the results obtained from fluorescence enhancement measurements. A remarkable increase in the fluorescence anisotropy values suggested that 3,7-diHF bound at a motional restricted pocket on HSA. The results indicated that 3,7-diHF, in anionic form, was bound within the hydrophobic pockets of the subdomain IIA of HSA (site I), and stabilised mainly by electrostatic force and ionic interactions. The binding mode of drug-protein was discussed based on above experimental results.
通过荧光猝灭、荧光增强、稳态和时间分辨荧光发射以及紫外可见吸收光谱法研究了3,7 - 二羟基黄酮（3,7 - diHF）与人血清白蛋白（HSA）的相互作用机制。通过分别研究3,7 - diHF - HSA复合物在pH 7.4和pH 3.5时的光谱性质来确定3,7 - diHF在蛋白质上的结合位点，并通过位点标记竞争实验进行了确认。通过荧光猝灭实验估算了3,7 - diHF - HSA复合物的结合参数，所得数据与荧光增强测量结果吻合良好。荧光各向异性值显著增加表明3,7 - diHF结合在HSA上一个运动受限的口袋处。结果表明，呈阴离子形式的3,7 - diHF结合在HSA亚结构域IIA的疏水口袋内（位点I），主要通过静电力和离子相互作用实现稳定。基于上述实验结果对药物 - 蛋白质的结合模式进行了讨论。
