光谱法和分子对接法研究和比较两种全氟羧酸与人血清白蛋白的结合模式（英文）

A New Strategy to Probe and Compare the Binding Modes of Two Perfluorocarboxylic Acids with Human Serum Albumin Based on Spectroscopic and Molecular Docking Methods

全氟羧酸(PFCAs)由于具有既亲水又疏水的表面活性剂特性,被广泛应用于工业和生活产品中。全氟十一酸(PFUnA)和全氟十三酸(PFTriA)是长链PFCAs类的典型代表,但近年来它们越来越频繁的在人体中检测到,并且发现表现出内分泌干扰效应、发育毒性和致畸性。本文以光谱学和分子对接为基础,探索PFUnA和PFTriA与人体最丰富的蛋白人血清白蛋白(HSA)的结合模式。结果表明,PFUnA和PFTriA均通过动静态猝灭过程猝灭HSA的内源荧光,与HSA只有一个强亲和位点,且PFUnA与HSA的结合比PFTriA更紧密。根据热力学计算结果,可知PFUnA与HSA结合的焓变、熵变分别为-26.32 kJ·mol^(-1)和21.76 J·mol^(-1)·K^(-1),其结合作用主要依靠静电引力,而PFTriA主要通过范德华力和卤键与HSA结合,是放热熵减过程,其焓变和熵变分别为-39.69 kJ·mol^(-1)和-25.66 J·mol^(-1)·K^(-1)。计算得到的结合距离(r<8 nm)显示从HSA到PFUnA和PFTriA发生了非辐射能量转移。三维荧光光谱和圆二色谱表明,PFUnA和PFTriA与HSA的结合不仅可以改变HSA的构象和微环境,还可以引起α-螺旋稳定性降低。取代实验和分子对接进一步显示PFUnA和PFTriA通过极性键、疏水作用力和卤键等与HSA的亚域ⅡA疏水腔有高亲和性,且荧光团Trp残基处于结合位置中,进一步证明PFUnA和PFTriA可以猝灭HSA的荧光。本文研究结果为阐明长链PFCAs在机体内与血清蛋白的结合机理提供了完整可靠的数据,并为长链PFCAs的毒性评价和毒理学研究提供了理论依据。

Perfluorocarboxylic acids（PFCAs） have been widespread used for over half century as surfactants in commercial and industrial products because of their hydrophilic and hydrophobic peculiarity.Perfluoroundecanoic acid（PFUnA） and perfluorotridecanoic acid（PFTriA） are two representatives of long-chain PFCAs,and they were detected more frequently in human body recent years,however,the two PFCAs were found to express endocrine disruption effects,developmental toxicity and teratogenicity.In this study,we established a new strategy to probe the binding modes of PFUnA（PFTriA） with the most abundant protein human serum albumin（HSA） based on spectroscopic and molecular docking methods.Results showed that both PFUnA and PFTriA can quench the intrinsic fluorescence of HSA with one binding site by means of dynamic and static quenching procedure with a strong affinity and the order is PFUnAPFTriA.On the basis of thermodynamic results,we knew that the main driving force of the interaction between PFUnA and HSA was electrostatic force（ΔH=-26.32 kJ · mol^-1,ΔS=21.76 J · mol^-1 · K^-1,while van der Waals interaction and halogenbond played major roles in complexation process of PFTriA-HSA（ΔH=-39.69 kJ · mol^-1,AS=-25.66 J · mol^-1· K ^-1.The binding distance（r8 nm） indicated that the non-radioactive energy transfer came into being from HSA to PFUnA（PFTriA）.The binding process of PFUnA（PFTriA） with HSA caused conformational and some micro-environmental changes of HSA,but also led to a loss of helical stability through three-dimensional fluorescence and circular dichroism spectra（CD）.Furthermore,site markers competitive experiments and molecular docking revealed that PFUnA and PFTriA had a high affinity into hydrophobic pocket of subdomain IIA in HSA through polar force,hydrophobic interaction and halogen-bond and so on,and the fluorophore Trp residues was located in the binding position which proved further the quenching of PFUnA and PFTriA on HSA fluorescence.The accurate and full basic data in the work are beneficial to clarify the binding mechanism of long-chain perfluorocarboxylic acids with serum protein in vivo,and provide essential theoretical clues for their toxicity assessment and toxicologic research.