Li Xiangrong, Wang Gongke, Chen Dejun, Lu Yan
School of Chemistry and Chemical Engineering, Key Laboratory of Green Chemical Media and Reactions, Ministry of Education, Henan Normal University, Xinxiang, Henan 453007, PR China.
Mol Biosyst. 2014 Feb;10(2):326-37. doi: 10.1039/c3mb70373h.
Binding of ascorbic acid (water-soluble antioxidant) and α-tocopherol (lipid-soluble antioxidant) to bovine serum albumin (BSA) has been studied using isothermal titration calorimetry (ITC), in combination with fluorescence spectroscopy, UV-vis absorption spectroscopy and Fourier transform infrared (FT-IR) spectroscopy. Thermodynamic investigations reveal that ascorbic acid/α-tocopherol binding to BSA is driven by favorable enthalpy and unfavorable entropy, and the major driving forces are hydrogen bonding and van der Waals forces. For ascorbic acid, the interaction is characterized by a high number of binding sites, which suggests that binding occurs by a surface adsorption mechanism that leads to coating of the protein surface. For α-tocopherol, one molecule of α-tocopherol combines with one molecule of BSA and no more α-tocopherol binding to BSA occurs at concentration ranges used in this study. Fluorescence experiments suggest that ascorbic acid has predominantly a "sphere of action" quenching mechanism, whereas, for α-tocopherol, the quenching mechanism is "static quenching" and due to the formation of a ground state complex. Additionally, as shown by the UV-vis absorption, synchronous fluorescence spectroscopy, and FT-IR, ascorbic acid and α-tocopherol may induce conformational and microenvironmental changes of BSA.
已采用等温滴定量热法（ITC），结合荧光光谱、紫外可见吸收光谱和傅里叶变换红外（FT-IR）光谱，研究了抗坏血酸（水溶性抗氧化剂）和α-生育酚（脂溶性抗氧化剂）与牛血清白蛋白（BSA）的结合情况。热力学研究表明，抗坏血酸/α-生育酚与BSA的结合由有利的焓和不利的熵驱动，主要驱动力是氢键和范德华力。对于抗坏血酸，这种相互作用的特点是有大量结合位点，这表明结合是通过表面吸附机制发生的，导致蛋白质表面被覆盖。对于α-生育酚，一分子α-生育酚与一分子BSA结合，在本研究使用的浓度范围内，没有更多的α-生育酚与BSA结合。荧光实验表明，抗坏血酸主要具有“作用域”猝灭机制，而对于α-生育酚，猝灭机制是“静态猝灭”，且是由于形成了基态复合物。此外，紫外可见吸收、同步荧光光谱和FT-IR结果表明，抗坏血酸和α-生育酚可能会诱导BSA的构象和微环境变化。
