静电和疏水效应对TGEV主蛋白酶二聚体稳定性的影响

从TGEV 3CL蛋白酶二聚体结构出发,研究了TGEV 3CL蛋白酶二聚体单体之间的静电和疏水相互作用.蛋白质的静电相互作用通过有限差分方法求解Poisson-Boltzmann方程得到,疏水相互作用通过分析溶剂可及性表面模型得到.考察了不同pH值对TGEV 3CL蛋白酶二聚体静电和疏水相互作用的影响,在pH值为5.5～8.5时,二聚体静电相互作用能、静电去溶剂化能和疏水自由能都较小,表明在该条件下静电和疏水相互作用有利于二聚体的稳定存在,这符合实验结晶所需条件.pH值对静电去溶剂化能的影响大于疏水自由能,表明静电作用是造成强酸或强碱条件下二聚体不能稳定存在的主要原因.     

SARS冠状病毒主要蛋白酶二聚体静电和疏水效应的研究

从三种冠状病毒主要蛋白酶SARS 3CL, HCoV 3CL和TGEC 3CL蛋白酶结构出发,着重研究了三种蛋白酶二聚体单体之间的静电和疏水相互作用.用连续介质模型有限差分方法计算得到三种蛋白二聚体界面处的静电势,发现三种蛋白酶单体和单体之间静电势分布具有明显的互补性,三种蛋白酶二聚体单体之间具有相同的静电相互作用能.用溶剂可及表面积模型分析了分子表面积及疏水性,发现三种蛋白酶具有相同的疏水分布,其中SARS 3CL蛋白酶疏水率为74%,驱动其单体聚合成二聚体.对三种蛋白酶的去溶剂化能疏水项的计算表明,三种蛋白酶二聚体单体之间具有相似的疏水相互作用能.     

SARS冠状病毒3CL蛋白酶及其抑制剂的理论研究

应用AutoDock程序将SARS冠状病毒3CL蛋白酶及其抑制剂配体和受体进行了对接,并用InsightⅡ中的Discover 3模块进行了分子动力学模拟,分析了蛋白酶活性口袋的形状,讨论了其亚基的氢键、静电、疏水等相互作用,为进一步设计药物提供了重要的参考信息.     

SARS 3CL蛋白酶同源二聚化与底物结合互为别构调控因素

研究了严重急性呼吸系统综合症(SARS)冠状病毒3C-Like蛋白酶(3CLpro)在存在底物或抑制剂时的二聚体形成情况. 通过测定酶活性随酶浓度的变化, 拟合出在底物存在下酶二聚体的解离常数约为0.94 μmol·L-1, 小于纯蛋白酶的二聚体解离常数(14.0 μmol·L-1), 表明底物对二聚体的形成具有增强作用. 选用与底物具有类似结合方式的靛红类抑制剂N-萘甲基靛红-5-甲酰胺(5f), 利用超速离心沉降速率方法定量测定了SARS 3CL蛋白酶单体和二聚体在不同浓度5f时的含量, 发现5f同样具有诱导二聚体形成的能力. 在3 μmol·L-1蛋白酶浓度下测定得到诱导二聚的EC50 值(半数有效浓度)约为1 μmol·L-1, 说明二聚体中只有一个单体与抑制剂结合. 研究结果表明, 随着底物浓度的升高, SARS 3CL蛋白酶会形成更多的二聚体, 而二聚体含量的提高又反过来提高酶的活性, 这种双向别构调控机制有可能是病毒用来调控多聚蛋白水解速率和组装时机的一种方法.     

非肽类SARS冠状病毒3CL蛋白酶抑制剂的设计与活性表征

SARS冠状病毒3CL蛋白酶是SARS病毒复制过程中的主要蛋白酶, 针对其开展药物设计有望得到有效的抗SARS病毒药物. 本文基于SARS冠状病毒3CL蛋白酶的三维结构, 对现有化学试剂及临床用药数据库进行虚拟筛选, 选出可能对SARS冠状病毒3CL蛋白酶有抑制的非肽化合物进行初步活性测试, 并研究了已知的人鼻病毒3C蛋白酶抑制剂对SARS冠状病毒3CL蛋白酶的活性, 合成了两种母环的衍生物, 得到靛红和哌嗪两类SARS冠状病毒3CL蛋白酶的抑制剂, 其中一个靛红类化合物的IC₅₀为0.76 µmol•L^－1; 而抗组胺药哌嗪类化合物对SARS冠状病毒3CL蛋白酶及细胞培育的SARS病毒的抑制作用, 提示了老药可以开发出新的用途.     

苯、甲苯、联苯在水-叔丁醇混合溶剂中的溶解度和疏水作用

在283.15,288.15,293.15,298.15 K温度下,测定了苯、甲苯、联苯在水-叔丁醇混合溶剂中的溶解度,其中混合溶剂中叔丁醇的物质的量分数分别为0.000,0.010,0.020,0.030,0.040,0.045,0.050,0.060,0.080,0.100. 以此为基础计算了苯、甲苯、联苯在不同温度和溶剂组成条件下的标准溶解Gibbs自由能和相关过程的疏水相互作用Gibbs自由能,并从微观上讨论了溶剂结构对溶解度和疏水作用的影响.     

固体的表面能及其亲水/疏水性

固体表面的亲水/疏水性质与表面分子与水之间的固/液界面相互作用自由能以及水介质中表面分子与空气之间的固/(液)/气界面相互作用自由能密切相关.水介质中固体表面与水之间存在范德华引力或疏水引力,与气泡之间存在范德华斥力、疏水引力以及静电斥力.在Lifshitz-范德华(LW)相互作用自由能、Lewis酸-碱(AB)相互作用自由能以及静电(EL)相互作用自由能3者之中,AB相互作用自由能比其它两者要大2～3个数量级以上.与固体表面能Lewis酸-碱分量相关的亲水指数√r_s~++√r_s~-可以成为衡量固体表面亲水/疏水性质的重要判据,水介质中固体表面疏水的必要条件是√r_s~++√r_s~-＜5mJ~(1/2)/m,指数大于或接近5mJ~(1/2)/m的表面必然是亲水的.     

三种金属硫蛋白聚合物静电效应的研究

考察了三种金属硫蛋白(大鼠金属硫蛋白亚型II,兔肝金属硫蛋白亚型I和兔肝金属硫蛋白亚型II)的单体、二聚体和三聚体在pH为5.6-8.5和10.6两种缓冲条件下的静电势分布。其中大鼠金属硫蛋白亚型II的结构直接来自于晶体数据,兔肝金属硫蛋白亚型I和II的结构则通过同源蛋白模型搭建。三种金属硫蛋白的静电势通过有限差分方法求解Poisson-Boltzmann方程得到。对于三种金属硫蛋白的二聚体,pH为5.6-8.5时,单体和单体之间的静电势分布具有明显的互补性;但pH≥10.6时,这种互补性会大大削弱。对于三种金属硫蛋白的三聚体,单体和二聚体之间主要表现为静电排斥,而且pH在10.6下的静电排斥力明显强于pH为5.6-8.5时的静电排斥。     

新型SARS-CoV 3CL蛋白酶荧光多肽底物的设计、制备及其酶动力学研究

以邻氨基苯甲酸（Abz）为荧光发射基团、2，4-二硝基苯基乙二胺（rdanp）为荧光猝灭基团，设计合成了SARS，CoV3CL蛋白酶的新型荧光多肽底物：H2N-E（Eddnp）STLQSGLK（Abz）-CONH2．用液相色谱-质谱（LC—MS）联用技术进行了表征，表明该多肽底物能被SAILS-CoV 3CL蛋白酶识别，并在QS之间被专一性酶解．另外，利用该多肽底物的荧光共振能量转移（FRET）特性，对SARS—CoV 3CL蛋白酶的酶解动力学性质进行了研究，结果表明，此荧光多肽底物可以作为荧光探针，应用于SARS—CoV 3CL蛋白酶活性的测定及其抑制剂的筛选．     

胺基化PGMA交联微球对胆红素的吸附机理

通过胺基与环氧键之间的开环反应, 用己二胺及多乙烯多胺等小分子胺化试剂对聚甲基丙烯酸缩水甘油酯(PGMA)交联微球进行了化学改性, 制得了胺基化的PGMA交联微球, 研究了该功能微球对胆红素的吸附特性, 考察了胺化试剂的分子结构、介质pH值、离子强度及温度等因素对其吸附性能的影响, 较深入地研究了吸附机理. 实验结果表明, 胺基化微球对胆红素具有强吸附作用, 吸附容量可达17.80 mg·g-1, 等温吸附服从Freundlich方程. 胺基化微球与胆红素分子之间的作用力以静电相互作用为主, 同时也存在氢键作用与疏水相互作用. 在pH 值为6 的介质中二者之间的静电作用最强, 胆红素吸附容量最高. 高离子强度不利于静电相互作用, 盐度增大使吸附容量减小. 温度升高有利于疏水相互作用而不利于氢键作用, 两种作用中占优势者主导温度对吸附容量的影响. 用己二胺改性的微球, 由于疏水相互作用的强化以及较长连接臂导致较小的空间位阻, 使其对胆红素的吸附能力明显高于多乙烯多胺改性的微球.     

Simple physics-based analytical formulas for the potentials of mean force for the interaction of amino acid side chains in water. 1. Approximate expression for the free energy of hydrophobic association based on a Gaussian-overlap model

A physics-based model is proposed to derive approximate analytical expressions for the cavity component of the free energy of hydrophobic association of spherical and spheroidal solutes in water. The model is based on the difference between the number and context of the water molecules in the hydration sphere of a hydrophobic dimer and of two isolated hydrophobic solutes. It is assumed that the water molecules touching the convex part of the molecular surface of the dimer and those in the hydration spheres of the monomers contribute equally to the free energy of solvation, and those touching the saddle part of the molecular surface of the dimer result in a more pronounced increase in free energy because of their more restricted mobility (entropy loss) and fewer favorable electrostatic interactions with other water molecules. The density of water in the hydration sphere around a single solute particle is approximated by the derivative of a Gaussian centered on the solute molecule with respect to its standard deviation. On the basis of this approximation, the number of water molecules in different parts of the hydration sphere of the dimer is expressed in terms of the first and the second mixed derivatives of the two Gaussians centered on the first and second solute molecules, respectively, with respect to the standard deviations of these Gaussians, and plausible analytical expressions for the cavity component of the hydrophobic-association energy of spherical and spheroidal solutes are introduced. As opposed to earlier hydration-shell models, our expressions reproduce the desolvation maxima in the potentials of mean force of pairs of nonpolar solutes in water, and their advantage over the models based on molecular-surface area is that they have continuous gradients in the coordinates of solute centers.     

Molecular origin of anticooperativity in hydrophobic association

The structuring of water molecules in the vicinity of nonpolar solutes is responsible for hydrophobic hydration and association thermodynamics in aqueous solutions. Here, we studied the potential of mean force (PMF) for the formation of a dimer and trimers of methane molecules in three specific configurations in explicit water to explain multibody effects in hydrophobic association on a molecular level. We analyzed the packing and orientation of water molecules in the vicinity of the solute to explain the effect of ordering of the water around nonpolar solutes on many-body interactions. Consistent with previous theoretical studies, we observed cooperativity, manifested as a reduction of the height of the desolvation barrier for the trimer in an isosceles triangle geometry, but for linear trimers, we observed only anticooperativity. A simple mechanistic picture of hydrophobic association is drawn. The free energy of hydrophobic association depends primarily on the difference in the number of water molecules in the first solvation shell of a cluster and that in the monomers of a cluster; this can be approximated by the molecular surface area. However, there are unfavorable electrostatic interactions between the water molecules from different parts of the solvation shell of a trimer because of their increased orientation induced by the nonpolar solute. These electrostatic interactions make an anticooperative contribution to the PMF, which is clearly manifested for the linear trimer where the multibody contribution due to changes in the molecular surface area is equal to zero. The information theory model of hydrophobic interactions of Hummer et al. also explains the anticooperativity of hydrophobic association of the linear trimers; however, it predicts anticooperativity with a qualitatively identical distance dependence for nonlinear trimers, which disagrees with the results of simulations.     

Complexation and chiral recognition thermodynamics of 6-amino-6-deoxy-beta-cyclodextrin with anionic, cationic, and neutral chiral guests: counterbalance between van der Waals and coulombic interactions

The stability constant (K), standard free energy (Delta G degrees), enthalpy (Delta H degrees), and entropy changes (T Delta S degrees) for the complexation of 6-amino-6-deoxy-beta-cyclodextrin with more than 50 negatively or positively charged as well as neutral guests, including 22 enantiomer pairs, have been determined in aqueous phosphate buffer (pH 6.9) at 298.15 K by titration microcalorimetry. The thermodynamic parameters obtained in this study and the relevant data for native beta-cyclodextrin indicate that the complexation and chiral discrimination behavior of the cationic host with charged guests are governed by the critical counterbalance between the electrostatic interactions of the charged groups in host and guest and the conventional intracavity interactions of the hydrophobic moiety of guest, such as hydrophobic, van der Waals, solvation/desolvation, and hydrogen-bonding interactions.     

Emission of thioflavin T and its off-on control in polymer membranes.

The absorption and emission spectral properties of thioflavin T (TFT+) in Nafion (Nf) and cellulose matrices have been studied. Formation of the emittive dimer is observed in both matrices. The monomer TFT+ emission is blueshifted in Nafion membrane (Nf), whereas it is red-shifted in cellulose membrane when compared with the emission in aqueous solution. The dimer emission of TFT+ in the Na+-Nf membrane undergoes off-on switching with acids and alkalis. The TFT+ molecule undergoes protonation in the H+-Nf and the protonated dye is fluorescent. The dimer emission of TFT+ is not observed in the dry H+-Nf membrane because of the protonation of the TFT+ molecule. The diffusion coefficient and the free energy of hydrophobic interaction for the TFT+ molecule in the Nf membrane are calculated. The TFT+ molecule experiences hydrophobic and electrostatic interactions in the Nf matrix, whereas it experiences a polar environment in the cellulose membrane. The 3D emission spectral studies support the formation of the emittive dimer in both Nf and cellulose matrices.     

Thermodynamics of molecular recognition of bile salts by 3,6'-(oligoethylenediamino-bridged) beta-cyclodextrin dimers

The molecular recognition behaviors of some representative bile salts by three 3,6'-bridged beta-cyclodextrin dimers with oligo(ethylenediamino) linkers in different lengths, i.e. 3,6'-(ethylenediamino-bridged) beta-cyclodextrin dimer (1), 3,6'-(diethylenetriamino-bridged) beta-cyclodextrin dimer (2), and 3,6'-(triethylenetetraamino-bridged) beta-cyclodextrin dimer (3), were investigated in aqueous phosphate buffer solution (pH 7.20) at 25 degrees C by means of 2D NMR spectroscopy and isothermal titration microcalorimetry. Owing to the cooperative host-linker-guest binding mode between host and guest, these 3,6'-bridged beta-cyclodextrin dimers showed significantly enhanced binding abilities and molecular selectivities as compared with native beta-cyclodextrin through the simultaneous contributions of hydrophobic, hydrogen bond, and electrostatic interactions. Thermodynamically, the inclusion complexations of these beta-cyclodextrin dimers with bile salts were mainly driven by large enthalpic gain, accompanied by slight to moderate entropic loss. An enthalpy-entropy compensation analysis demonstrated that these beta-cyclodextrin dimers experienced large conformational changes and extensive desolvation effect upon inclusion complexation with guest molecules.     

胺类分子结构对油酸囊泡pH窗口的影响

基于激光丁达尔效应及浊度测定, 考察了改变二元胺的碳桥长度、 多元胺的氨基多寡、 长链伯胺的碳链长度及季铵化等因素对油酸囊泡pH窗口的影响. 结果表明, 二元胺及多元胺主要导致油酸囊泡的pH窗口向碱性方向拓宽, 而长链伯胺和十二烷基三甲基溴化铵(DTAB)不仅可使油酸囊泡的pH窗口向碱性拓宽, 而且可使油酸在酸性pH区域形成另一个介稳至稳定的新囊泡相. 氨基在不同pH下质子化和脱质子化转换是胺类分子调节油酸囊泡pH窗口的共同驱动力, 疏水作用是长链胺类的又一驱动力, 而静电吸引是季铵盐的另一种特殊驱动力. 分子间相互作用的热力学参数及结合能计算结果表明, 二乙烯三胺为代表的二元胺或多元胺在油酸囊泡表面以氢键或离子-偶极作用等非共价结合为主, 其调节功能弱于长链伯胺及DTAB与油酸的疏水共组装或DTAB与油酸的静电吸引作用.     

DNA-surfactant interactions: a procedure for determination group contributions

A procedure, based on the measurement of pyren-1-carboxyaldehyde fluorescence, has been used to obtain the free energy corresponding to the interaction of several surfactants of alkyltrimethylammonium-type RN(CH3)3+ and DNA. These free energies depend linearly on the number of carbon atoms in the tail of the surfactant. In this way, it was possible to determine the -CH2-/DNA interaction free energy as well as the free energy of interaction of DNA and the head group of the surfactants. According to the values of these free energies, the surfactant/DNA interaction is mainly electrostatic, for the surfactants used in this work. However, for long enough tails the free energy corresponding to the hydrophobic interaction can reach the same value as the electrostatic one. The procedure used in this work could be extended to the measurement of other group/DNA interactions.     

pH响应木质素基胶体球的制备和表征

以造纸制浆废液中的松木碱木质素（AL）为原料,通过季铵化改性,制备了季铵化碱木质素（QAL）.QAL与十二烷基苯磺酸钠（SDBS）通过静电作用形成QAL/SDBS复配物,将QAL/SDBS复配物在乙醇/水混合溶剂中进行自组装得到具有pH响应性的胶体球.采用X射线光电子能谱（XPS）、扫描电子显微镜（SEM）、透射电子显微镜（TEM）、元素分析和静态接触角研究了胶体球的形成过程和结构.研究结果表明,QAL/SDBS复配物通过疏水聚集作用形成具有较疏水的“核”和较亲水的“壳”结构的规整胶体球.在pH=3.0时,由于QAL与SDBS间的静电作用和疏水作用使胶体球能够稳定存在.当pH>7.5时,季铵化碱木质素上的羧基电离,由于静电斥力的作用使胶体球开始解聚,当pH=10.5时,季铵化碱木质素上的酚羟基的电离使得QAL与SDBS间的静电斥力增大,胶体球完全解聚.这种在酸性条件下稳定,中性条件下解聚的胶体球在药物缓释方面具有潜在的应用.