Department of Biology, Faculty of Sciences, Mashhad Branch, Islamic Azad University, Mashhad, Iran.
J Biomol Struct Dyn. 2011 Aug;29(1):181-206. doi: 10.1080/07391102.2011.10507382.
The interaction between cyclophosphamide hydrochloride (CYC) and aspirin (ASA) with human serum albumin (HSA) was studied by various kind of spectroscopic, ζ potential and molecular modeling under physiological conditions. The fluorescence data showed that the binding of drugs to proteins caused strong static fluorescence quenching. The analysis of the fluorescence quenching of HSA in the binary and ternary systems displayed that ASA was affected by the complex formed between CYC and HSA. Moreover, CYC was influenced by the HSA-ASA complex. The inherent binding information, including the quenching mechanism, binding constants, number of binding sites, effective quenching constant, fraction of the initial fluorescence and thermodynamic parameters were measured by the fluorescence quenching technique at various temperatures. In addition, according to the synchronous fluorescence spectra of HSA, the results showed that the fluorescence quenching of HSA originated from the Trp and Tyr residues, and indicated a conformational change of HSA with the addition of the drugs. Far-UV CD spectra of HSA were recorded before and after the addition of ASA and CYC as binary and ternary systems. An increase in intensity of the positive CD peak of HSA was observed in the presence of the drugs. The results were interpreted by excited interactions between the aromatic residues of the HSA binding sites and the drugs bound to them. The distance r between donor and acceptor was obtained by the Forster energy according to fluorescence resonance energy transfer (FRET) and found to be 2.35 nm and 1.78 nm for CYC and ASA, respectively. This confirmed the existence of static quenching for proteins in the presence of CYC and ASA. Furthermore, docking studies pointed at a reduction of the affinity of each of the drug compounds to the protein in the presence of the other in meaningful amounts. Pre-binding of any of the said compounds forced the second to bind in a non-optimized location and orientation. The potential at the electrokinetic shear surface of the protein-drug solution were measured at several concentrations of the drugs by the ζ potential technique, which confirmed experimental and theoretical results.
在生理条件下，通过各种光谱、ζ 电位和分子模拟方法研究了盐酸环磷酰胺（CYC）和阿司匹林（ASA）与人血清白蛋白（HSA）的相互作用。荧光数据表明，药物与蛋白质的结合导致强静态荧光猝灭。二元和三元体系中 HSA 荧光猝灭的分析表明，ASA 受到 CYC 和 HSA 形成的复合物的影响。此外，CYC 受到 HSA-ASA 复合物的影响。通过荧光猝灭技术在不同温度下测量了包括猝灭机制、结合常数、结合位点数、有效猝灭常数、初始荧光分数和热力学参数在内的固有结合信息。此外，根据 HSA 的同步荧光光谱，结果表明 HSA 的荧光猝灭源于色氨酸和酪氨酸残基，表明随着药物的加入，HSA 的构象发生了变化。在加入 ASA 和 CYC 作为二元和三元体系前后记录了 HSA 的远紫外 CD 光谱。在药物存在的情况下，观察到 HSA 的正 CD 峰强度增加。通过芳香族残基与结合到它们上的药物之间的激发相互作用解释了结果。根据荧光共振能量转移（FRET），通过 Förster 能量获得供体和受体之间的距离 r，对于 CYC 和 ASA，分别为 2.35nm 和 1.78nm。这证实了在存在 CYC 和 ASA 的情况下蛋白质存在静态猝灭。此外，对接研究表明，在存在大量其他药物化合物的情况下，每种药物化合物与蛋白质的亲和力都会降低。任何一种上述化合物的预结合都会迫使第二种以非最佳位置和方向结合。通过 ζ 电位技术在药物的几个浓度下测量蛋白质-药物溶液的电动剪切表面的电位，证实了实验和理论结果。
