Zhang Yaheng, Yue Yuanyuan, Li Jiazhong, Chen Xingguo
Department of Chemistry, Lanzhou University, Lanzhou 730000, China.
J Photochem Photobiol B. 2008 Mar 28;90(3):141-51. doi: 10.1016/j.jphotobiol.2007.12.004. Epub 2007 Dec 31.
In this work, fluorescence quenching technique, Fourier transform infrared (FT-IR) spectroscopy, circular dichroism (CD) spectroscopy and dynamic light scattering (DLS) technique were used to gain the binding information of caffeic acid and human serum albumin (HSA) in AOT/isooctane/water microemulsions. The interaction of HSA with caffeic acid at 296, 303, and 310 K in omega(0) 20 microemulsions was characterized by one binding site with the affinity constant K at (3.23+/-0.01) x 10(4), (3.06+/-0.03) x 10(4) and (2.82+/-0.05) x 10(4)M(-1), respectively. The affinities in microemulsions are much higher than that in buffer solution. The CD spectra and FT-IR spectra with qualitative and quantitative results proved that the protein secondary structure changed in the microemulsions in the absence and presence of caffeic acid compared with the free form of HSA in buffer. The binding process was exothermic and spontaneous, as indicated by the thermodynamic analyses. These data indicated that hydrophobic interaction played a major role in the binding of caffeic acid to HSA in microemulsions and electrostatic interaction can not be excluded. The displacement experiments confirmed that caffeic acid could bind to the site I of HSA, which was in agreement with the result of the molecular modeling study. Furthermore, the DLS data suggested that HSA may locate at the interface of the microemulsion and caffeic acid could interact with them.
在本研究中，采用荧光猝灭技术、傅里叶变换红外（FT-IR）光谱、圆二色（CD）光谱和动态光散射（DLS）技术来获取咖啡酸与AOT/异辛烷/水微乳液中人类血清白蛋白（HSA）的结合信息。在ω(0) 20微乳液中，296、303和310 K下HSA与咖啡酸的相互作用以一个结合位点为特征，亲和常数K分别为(3.23±0.01)×10⁴、(3.06±0.03)×10⁴和(2.82±0.05)×10⁴ M⁻¹。微乳液中的亲和力远高于缓冲溶液中的亲和力。CD光谱和FT-IR光谱的定性和定量结果证明，与缓冲液中游离形式的HSA相比，在有无咖啡酸存在的情况下，微乳液中蛋白质的二级结构发生了变化。热力学分析表明，结合过程是放热且自发的。这些数据表明，疏水相互作用在微乳液中咖啡酸与HSA的结合中起主要作用，静电相互作用也不能排除。置换实验证实咖啡酸可与HSA的位点I结合，这与分子模拟研究的结果一致。此外，DLS数据表明HSA可能位于微乳液的界面，且咖啡酸可与其相互作用。
