摘要
在生理酸度(pH 7.4)条件下,采用荧光光谱、紫外-可见吸收光谱、圆二色谱(CD)和红外光谱(FT-IR)等多种光谱方法并结合分子模拟技术,测定了塑化剂邻苯二甲酸二正辛酯(DnOP)与人血清白蛋白(HSA)的相互作用模式。荧光滴定结果表明,DnOP对HSA内源荧光的猝灭机制为形成HSA-DnOP复合物的静态猝灭,其在不同温度下的熵变(ΔS°)和焓变(ΔH°)分别为35.32 J·mol-1·K-1和-9.13 kJ·mol-1,表明结合反应主要由疏水作用和氢键驱动。位点竞争实验表明DnOP与曙红Y发生了置换反应,揭示DnOP主要结合在HSA亚结构域ⅡA(SiteⅠ位),分子模拟结果显示,DnOP插入亚结构域ⅡA的疏水空腔,通过疏水作用以及DnOP的羰基氧与His242氨基酸残基间形成的氢键与蛋白结合,实验结果与荧光光谱及位点竞争实验一致。紫外-可见光谱、CD及FT-IR光谱的分析结果表明,DnOP与HSA结合导致了HSA二级结构发生变化,降低了HSA中α-螺旋的含量,并诱导HSA的多肽链发生部分伸展。
The interaction mode of Di-n-octyl phthalate（ DnOP） with human serum albumin（ HSA） in physiological buffer（ pH 7. 4） was determined by multispectroscopic methods including fluorescence,Ultraviolet- visible（ UV- vis） absorption,circular dichroism（ CD） and Fourier transform infrared（ FT- IR） spectroscopy,coupled with molecular modeling technique. Results obtained from the analysis of fluorescence titration indicated that the fluorescence quenching of DnOP for HSA was a static procedure resulting. The thermodynamic analysis of the binding data obtained at different temperatures indicated that the interaction between DnOP and HSA was mainly driven by hydrophobic interactions and hydrogen bonds,as the values of the entropy change（ ΔS°） and the enthalpy change（ ΔH°） were found to be 35. 32 J·mol- 1·K- 1and- 9. 13 kJ·mol- 1,respectively. The site marker competitive experiments using different kinds of probes suggested a displacement reaction occurred between Eosin Y and DnOP with HSA. This result indicated that the binding site of DnOP to HSA lo- cated in the subdomain ⅡA（ Site Ⅰ）. Molecular modeling studies also proved that DnOP molecules inserted into the large hydrophobic activity of subdomainⅡA by the hydrophobic interactions and hydrogen bond between oxygen atom of carboxide and residue His242. UV- vis absorption,CD and FT- IR spectra revealed that the binding of DnOP to HSA changed the second structure of HSA with a loss of α-helical content,which suggested a partial unfolding of the polypeptide chain of the protein in the presence of DnOP.
出处
《分析测试学报》
CAS
CSCD
北大核心
2013年第12期1433-1437,共5页
Journal of Instrumental Analysis
基金
国家自然科学基金项目(31060210
21167013)
食品科学与技术国家重点实验室基金项目(SKLF-ZZB-201305
SKLF ZZA-201302
SKLF-KF-201203)
