He Ling-Ling, Wang Yong-Xia, Wu Xiao-Xia, Liu Xian-Ping, Wang Xin, Liu Bin, Wang Xin
College of Applied Chemistry, Shenyang University of Chemical Technology, Shenyang, 110142, China.
School of Pharmaceutical Sciences, Liaoning University, Shenyang, 110036, China.
Luminescence. 2015 Dec;30(8):1380-8. doi: 10.1002/bio.2910. Epub 2015 Mar 31.
In this work, the binding characteristics of methylene blue (MB) to human serum albumin (HSA) and the influence of Cu(2+) and Fe(3+) on the binding affinity of MB to HSA were investigated using fluorescence, absorption, circular dichroism (CD) spectroscopy and molecular modelling. The results of competitive binding experiments using the site probes ketoprofen and ibuprofen as specific markers suggested that MB was located in site I within sub-domain IIA of HSA. The molecular modelling results agreed with the results of competitive site marker experiments and the results of CD spectra indicated that the interaction between MB and HSA caused the conformational changes in HSA. The binding affinity of MB to HSA was enhanced but to a different extent in the presence of Cu(2+) and Fe(3+), respectively, which indicated that the influence of different metal ions varied. Enhancement of the binding affinity of MB to HSA in the presence of Cu(2+) is due to the formation of Cu(2+)-HSA complex leading to the conformational changes in HSA, whereas in the presence of Fe(3+), enhancement of the binding affinity is due to the greater stability of the Fe(3+)-HSA-MB complex compared with the MB-HSA complex.
在本研究中，利用荧光光谱、吸收光谱、圆二色光谱（CD）以及分子模拟等方法，研究了亚甲基蓝（MB）与人血清白蛋白（HSA）的结合特性，以及Cu(2+)和Fe(3+)对MB与HSA结合亲和力的影响。使用位点探针酮洛芬和布洛芬作为特异性标记物的竞争性结合实验结果表明，MB位于HSA亚结构域IIA内的位点I。分子模拟结果与竞争性位点标记实验结果一致，CD光谱结果表明MB与HSA之间的相互作用导致了HSA的构象变化。在分别存在Cu(2+)和Fe(3+)的情况下，MB与HSA的结合亲和力均增强，但增强程度不同，这表明不同金属离子的影响存在差异。在Cu(2+)存在下MB与HSA结合亲和力的增强是由于形成了Cu(2+)-HSA复合物，导致HSA构象发生变化，而在Fe(3+)存在下，结合亲和力的增强是由于Fe(3+)-HSA-MB复合物比MB-HSA复合物具有更高的稳定性。
