Chemical Sciences Division, Saha Institute of Nuclear Physics, Kolkata, India.
J Photochem Photobiol B. 2012 Mar 1;108:23-33. doi: 10.1016/j.jphotobiol.2011.12.005. Epub 2011 Dec 29.
Photophysical studies on binding interactions of a negatively charged anti-tumor photosensitizer, Merocyanine 540 (MC 540), with serum proteins, bovine serum albumin (BSA) and human serum albumin (HSA), have been performed using absorption and steady-state as well as time-resolved fluorescence techniques. Formation of ground state complex has been confirmed from the detailed studies of absorption spectra of MC 540 in presence of SAs producing isosbestic points. Binding between the proteins and MC 540, which perturbs the existing equilibrium between the fluorescent monomer and its non-fluorescent dimer, induces a remarkable enhancement in fluorescence anisotropy and intensity of MC 540 along with a red shift of its maximum. The binding stoichiometry of MC 540 and SAs are more than 1.0 which depicts that two types of complexes, i.e., 1:1 and 2:1 are formed with addition of varied concentration of protein. Both the steady-state and time-resolved fluorescence results show that in 2:1 complex one of the MC 540 molecules is exposed towards aqueous environment with a greater extent when bound with HSA compared to BSA due to the structural flexibility of that protein. Thermodynamic analyses using van't Hoff plot indicate that the binding between MC 540 and individual SA is an entropy-driven phenomenon. The probable hydrophobic binding site has been located by denaturation of proteins, micropolarity measurement and Förster resonance energy transfer and that is further supported by molecular docking studies. Changes in circular dichroism spectra of BSA in presence of MC 540 depict secondary structural changes of the protein. The induced-CD shows that BSA due to its rigid structure generates chirality in MC 540 much more efficiently compared to HSA.
采用吸收光谱、稳态和时间分辨荧光技术研究了带负电荷的抗肿瘤光敏剂 Merocyanine 540（MC 540）与血清蛋白牛血清白蛋白（BSA）和人血清白蛋白（HSA）之间的结合相互作用的光物理性质。通过详细研究 MC 540 在存在 SA 时的吸收光谱，产生等色点，证实了基态复合物的形成。MC 540 与蛋白质之间的结合，扰乱了荧光单体与其非荧光二聚体之间的现有平衡，导致 MC 540 的荧光各向异性和强度显著增强，并伴有最大波长红移。MC 540 和 SA 的结合化学计量数大于 1.0，这表明随着蛋白质浓度的变化，形成了两种类型的复合物，即 1:1 和 2:1。稳态和时间分辨荧光结果均表明，在 2:1 复合物中，与 BSA 相比，由于该蛋白质的结构灵活性，当与 HSA 结合时，MC 540 的一个分子更充分地暴露于水相环境中。使用范特霍夫图进行热力学分析表明，MC 540 与各别 SA 之间的结合是熵驱动的现象。通过蛋白质变性、微极性测量和Förster 共振能量转移定位了可能的疏水性结合位点，分子对接研究进一步支持了这一点。MC 540 存在下 BSA 的圆二色谱谱的变化表明了蛋白质的二级结构变化。诱导 CD 表明，与 HSA 相比，由于其刚性结构，BSA 更有效地在 MC 540 中产生手性。
