抗原肽与MHC分子相互作用QM/MM分子动力学模拟研究

在MHC I类(major histocompatibility complex class I)分子抗原加工提呈过程中抗原蛋白在抗原提呈细胞(antigen-presenting cells, APC)的胞浆中被蛋白酶体(proteasome)裂解成短肽peptide, 由转运相关蛋白(transporter associated with antigen processing, TAP)将蛋白酶体裂解产生的短肽片段从胞浆转运至内质网腔. 短肽peptide在内质网中与新生成的MHC I类分子结合, 形成peptide-MHC复合体被提呈到APC细胞表面, 与T细胞表面抗原受体(T cell receptor, TCR)特异性识别结合, 使得CTL细胞开始活化、增殖、分化, 进而对肿瘤细胞进行特异性杀伤. 目前对CTL细胞如何识别抗原肽-MHC复合物分子及抗原短肽peptide如何与主要组织相容性复合体MHC分子的相互作用识别结合的机理还不是很清楚. 传统的预测CTL细胞表位的方法没有考虑受体与配体结合过程中电子结构的变化, 电子结构的变化需要用量子力学方法来处理. 本文采用QM/MM多尺度生物大分子的分子动力学模拟方法, 以天然抗原肽TAX (LLFGYPVYVYU)与HLA-A*0201分子结合的晶体结构为模板, 替换抗原肽“锚点”氨基酸, 将口袋氨基酸残基的原子极化电荷在空间形成的静电势用电多极矩分量表示. 用箱线图分析每个口袋氨基酸分子静电势变化和功能, 确定Pocket B的Glu63和Lys66的功能是精细识别氨基酸和一级结合氨基酸, Pocket F的Asp77, Tyr84的功能是精细识别氨基酸, 而Asp77, Lys146是一级结合氨基酸, 表明QM/MM方法在提取抗原肽与MHC I类分子识别结合特异性信息是可行的, 这对了解免疫识别机理和指导肿瘤疫苗的开发都具有指导意义.     

应用独立成分分析方法预测CTL表位

基于独立成分分析方法分别采用3 z-scale和5 z-scale氨基酸结构描述符, 建立了抗原肽与MHC分子(major histocompatibility complex, MHC)相互作用结合的定量构效关系模型. 该两个模型训练集样本数是316, 预测集样本数是786. 结果表明: 3 z-scale模型的预测准确度和AUC值分别为70.3%, 0.70; 5 z-scale模型的预测准确度和AUC值分别为70.9%, 0.79. 本文建立CTL表位预测模型对进一步了解抗原肽与MHC I类分子相互作用机理具有一定的帮助.     

基于环境的编码方法在预测HLA-A*0201结合多肽中的应用

T淋巴细胞对抗原的识别是产生与调节有效免疫应答的关键, T细胞只识别主要组织相容性复合物(MHC)呈递上来的抗原, 因此MHC与抗原多肽的结合就成为一系列免疫应答过程中基础的一环. 为了辅助疫苗设计, 多种机器学习技术已被普遍应用于MHC结合多肽的预报领域中. 本文以支持向量机(SVM)为手段, 以HLA-A*0201的实验数据集为对象, 对多种肽段编码方法形成的模型进行评价, 得到的AUC值的范围在0.932~0.936之间. 提出一种新的利用抗原多肽结合环境的编码方法, 使预报的AUC值提高到0.953. 对独立数据集进行建模预报, 同样证明环境编码模型的预报准确率高于传统编码方法的准确率.     

蛋白酶体对抗原蛋白酶切特异性的研究

在MHCI类分子结合的抗原表位加工呈递途径中,泛素-蛋白酶体系统对抗原蛋白的降解发挥着重要作用。为了进一步研究蛋白酶体的酶切特异性,采用偏最小二乘方法（partial least squares method,PLS）建立了蛋白酶体的酶切位点预测模型,预测准确度为82.8％;由样本数据相应氨基酸对酶切位点形成的权重系数,得出蛋白酶体酶切位点及其两侧区域氨基酸的裂解特异性,它反映了蛋白酶体对抗原蛋白酶切的相互作用信息,也表明蛋白酶体对抗原蛋白酶切处理不是随机的,而是有一定模式和选择性的。     

多肽序列的结构特征与其MHC限制性

从多肽序列的一级结构出发,基于多肽序列中氨基酸侧链间距离和氨基酸侧链的电性特征,构建了多肽序列特征矢量,简称ζ矢量,选取文献中主要组织相容性复合物(MHC)中的14个A^d限制性和14个非A^d限制性多肽序列作为训练集建立辅助性T细胞(helperTlymphocytc,Th)表位预测的定量模型,为了检验该预报系统的精确性,进行了随机抽样检验和交互检验.结果表明,该预报系统具有稳定性好、预测能力强的特点,该方法可用于人的MHCⅠ和Ⅱ类表位的预测、蛋白质抗原免疫识别、亚单位疫苗分子设计及研制.     

基于原子力显微镜的人外周血CD8+T细胞形貌观察与力谱分析

体外分离人外周血CD8+T细胞,用植物凝结素(PHA)刺激活化,利用原子力显微镜(AFM)观察CD8+T细胞的形貌,并结合针尖修饰技术对力谱进行分析,确定了CD8抗原分子的分布.结果表明,与静息的CD8+T细胞相比,活化后的CD8+T细胞直径和高度增大,细胞表面变得更粗糙;CD8抗原-抗体的相互作用力大约是非特异性黏附力的5倍,活化后的CD8+T细胞上特异性黏附力(即CD8抗原分子位置)分布不均匀,其表面的CD8抗原分子分布以纳米团簇分布为主,CD8分子聚集更为明显,CD8+T细胞上CD8抗原-抗体相互作用力不随着活化而发生明显变化,说明CD8抗原-抗体之间具有高选择性.原子力显微镜为特异性T细胞的抗原识别和活化研究提供了一种新手段,能够使T细胞抗原识别和活化的机制得到更好地阐明.     

肽类树状大分子的合成及其在药物传输系统中的应用

树状大分子是近年来蓬勃发展的一类新型高分子材料, 其表面存在大量的官能团, 分子内部存在空腔且分子尺寸可控, 因此, 树状大分子已被广泛应用于众多的领域. 肽类树状大分子是指在树状大分子结构中含有肽键的一类大分子, 因其具有类似蛋白质一样的球状结构, 且具有优异的水溶性、生物相容性、生物降解性和细胞低毒性等特点, 所以, 肽类树状大分子可以作为药物传输的载体. 此外, 肽类树状大分子的疏水空腔可以装载疏水性药物, 对其起到增溶和缓释作用. 综述了肽类树状大分子的合成方法, 并对其与药物分子的结合机制及其在药物传输系统中的应用进行了总结与展望.     

基于金纳米颗粒的比色传感体系用于前列腺特异性膜抗原的检测

发展了一种基于金纳米颗粒的比色传感体系用于检测前列腺特异性膜抗原的新方法.实验中合成了带有正电荷的金纳米颗粒,并设计了一段带有负电荷的前列腺特异性膜抗原的底物肽段.该方法基于金纳米颗粒聚集状态不同导致颜色变化的性质以及酶与底物的特异性识别作用,达到前列腺特异性膜抗原的检测.带正电荷的金纳米颗粒与带负电的肽段产生静电相互作用,引起金纳米颗粒的聚集;当体系中加入前列腺特异性膜抗原后,由于前列腺特异性膜抗原与肽段的特异性识别作用,带负电的肽段水解为谷氨酸碎片分子,导致金纳米颗粒的分散,反应体系颜色变化快速、明显.该方法简单、灵敏,线性范围为2～10 nmol/L,检测限为0.5 nmol/L.此外,该方法可用于标准加入法测定尿液中的PSMA.     

毛细管电泳结合电喷雾质谱分析人类癌基因c-myb四链体核酸与活性天然产物的相互作用

药物与靶点间的作用关系直接影响到药理和药效。药物-靶点结合能力、结合计量关系等信息是药物研发过程中必需的表征数据。人类癌基因c-myb在结直肠癌等多种癌症组织中存在过度表达,目前已成为结直肠癌、白血病等癌症疾病潜在的治疗靶点。位于癌基因c-myb启动子区的一段富含鸟嘌呤（G）的DNA序列,通过阳离子的诱导可自发折叠形成分子内G-四链体,而小分子的特异性识别可以稳定该G-四链体,进而调节基因的转录和表达过程。该文采用压力辅助毛细管电泳前沿分析（PACE-FA）结合电喷雾质谱（ESI-MS）研究人类癌基因c-myb启动子G-四链体（G4）与天然产物分子间的相互作用。PACE-FA法在毛细管电泳前沿分析（CE-FA）过程中施加一个与分析物迁移同向的压力,在保证结果准确度的前提下,能够大大加快分析速度。同时结合ESI-MS,可快速解析结合分子与靶点的亲合力和化学计量关系。首先,利用ESI-MS快速筛选出3种有亲合力的天然产物,亲合力大小依次为：土荆皮乙酸>丁溴东莨菪碱>荷叶碱。考虑到溶液相中存在特异性与非特异性结合,接着用PACE-FA法准确分析溶液相中结合的特异性和结合常数。结果发现：丁溴东莨菪碱能够特异性结合靶点G4 DNA,结合比为1：1,结合常数为1.18×10⁵ L/mol;荷叶碱属于非特异性结合,而土荆皮乙酸并未与靶点G4 DNA形成复合物。该组合方法不仅分析速度快,而且能够提高亲和分析的准确度和特异性,有望应用于靶向药物先导结构的发现和作用机制评价。     

氯黄隆酶联免疫吸附分析技术研究

对邻氯苯磺酰胺进行衍生合成半抗原,并与载体蛋白质共价偶联制备突出氯黄隆分子特异性部分的合成抗原,以合成抗原免疫兔获得对氯黄隆具高亲合力的抗血清。采用硫酸铵盐析和ＤＥＡＥ纤维素反相吸附法分离纯化抗体,用辣根过氧化物酶以改良的过碘酶钠法标记 混合酸酐法标记半抗原。在此基础上建立了对氯黄隆具高度特异性的同接竞争,包被抗体,包被抗原直接竞争酶联免疫吸附分析技术。在优化条件下,氯黄隆测定的线性浓度范围为１０＾０－１０＾３ｎｇ／ｍＬ,检出限〈０．１ｎｇ／ｍＬ。邻氯苯磺酰胺及与氯黄隆结构类似的常用磺酰脲类除草剂不干扰氯黄隆的分析。     

二氧化碳和丙烯直接合成甲基丙烯酸的NiPMo/TiO2催化剂的研究

用溶胶-凝胶法以磷钼酸(MPA)的镍盐溶液水解钛酸四丁酯制备了NiPMo/TiO2催化剂.使用ICP、 XRD、 TG-DTA、 IR、 TPD-MS和微反应技术研究了催化剂的化学组成、热稳定性、化学吸附性质和催化反应性能.杂多钼酸盐与TiO2通过O2-在TiO2表面发生了键合.在623 K下,杂多阴离子仍保持原有的Keggin结构.CO2在Lewis酸位Ni(Ⅱ)和Lewis碱位Ni-O-Mo的桥氧协同作用下生成CO2卧式吸附态Ni(Ⅱ)←O-(CO)←(O--Ni).丙烯有多种吸附态在催化剂上吸附.在563 K、 1 MPa和空速1500 h-1的反应条件下,丙烯的摩尔转化率为3.2%,产物MAA选择性为95%.     

α-Amino acid derived enaminones and their application in the synthesis of N-protected methyl 5-substituted-4-hydroxypyrrole-3-carboxylates and other heterocycles

A new and simple synthesis of novel N-protected methyl 5-substituted-4-hydroxypyrrole-3-carboxylates, which exist in equilibrium with their 4-oxo tautomers, has been developed in two steps starting from N-protected α-amino acids. The key intermediates are enaminones, which can also be isolated, characterized, and used for the construction of other functionalized heterocycles, before they spontaneously decompose to pyrrole products. 4-Hydroxypyrroles are prone to partial aerial oxidation but can be efficiently alkylated or reduced to stable polysubstituted pyrrolidine derivatives.     

Specific intramolecular aromatic CH insertion of diazosulfonamides

The chemoselectivity in the intramolecular CH insertion of various diazosulfonamides has been experimentally studied. The results reveal that the aliphatic 1,4-, 1,5-, or 1,6-C(sp³)?H insertions of diazosulfonamides are not accessible, while the aromatic 1,5-C(sp²)?H insertion can be realized specifically by adjusting the diazo-adjacent group. In addition, the general chemoselectivities in the intramolecular CH insertions of diazosulfonyl compounds are summarized. Generally, diazosulfones undergo both aromatic 1,5-C(sp²)?H and aliphatic 1,5- and 1,6-C(sp³)?H insertions, while diazosulfonates undergo aliphatic 1,5- and 1,6-C(sp³)?H insertions. However, diazosulfonamides only undergo aromatic 1,5-C(sp²)?H insertion.     

N-Heterocyclic carbene-palladacyclic complexes: synthesis,characterization and their applications in the C-N coupling and α-arylation of ketones using aryl chlorides

N-Heterocyclic carbene-palladacyclic complexes 3 were successfully achieved in a one-pot procedure under mild conditions. The structure of 3a was unambiguously confirmed by X-ray single crystal diffraction and it was an active catalyst in the Buchwald-Hartwig amination and α-arylation of ketones even at very low catalyst loadings (0.01?mol%).     

Iodine-mediated oxidative Pictet-Spengler reaction using terminal alkyne as the 2-oxoaldehyde surrogate for the synthesis of 1-aroyl-β-carbolines and fused-nitrogen heterocycles

An efficient iodine-mediated oxidative Pictet-Spengler reaction in dimethyl sulphoxide (DMSO) using terminal alkynes as the 2-oxoaldehyde surrogate for the synthesis of aryl (9H-pyrido[3,4-b]indol-1-yl)methanones is described. The scope of the protocol includes the total synthesis of Fascaplysin, Eudistomins Y₁ and Y₂. The methodology is extended for preparing pyrrolo[1,2-a]-quinoxaline and indolo[1,5-a]quinoxaline derivatives. The utility of 1-aroyl-β-carbolines was demonstrated by performing palladium-catalyzed β-carboline directed ortho-C(sp2)-H functionalization of the phenyl ring with thiomethyl (SMe) group using DMSO as source and for accessing 4-aryl-canthin-6-ones.     

CoFe2O4自形成磁性液体场致结构化对磁化的影响

The Langevin paramagnetic theory can’t describe the relation between magnetization of ferrofluids and applied magnetic field. The structuralization of ferrofluids, which is considered the main influence factor of the magnetization, is regarded. The part of magnetization works is deposited when the structure is forming. This action influences the magnetization of ferrofluids directly or indirectly. On the base of the “compressing” model, the Langevin function that usually describes the magnetization of ferrofluid is modified, and a well-fitted curve is obtained. An equation of the relation between the equivalent volume fraction after being “compressed” and the intensity of magnetic field is discovered, which approximately describes the process of magnetization. The relation between the approximate initial susceptibility and the volume fraction can be obtained from modified formula.     

Facile synthesis of bicyclic 1-arylpyrazol-5-ones

In this Letter, we described a facile method for constructing fused bicyclic 1-arylpyrazol-5-one ring system. We employed various methylene-containing carboxylic acids as the substrates and proved that the pyrazolone ring closure requires activated methylene group in intermediate II. Accordingly, a series of structurally diversified, fused bicyclic 1-arylpyrazol-5-ones was prepared in moderate to high yields using the requisite substrates.     

Highly regioselective Buchwald–Hartwig amination at C-2 of 2,4-dichloropyridine enabling a novel approach to 2,4-bisanilinopyridine (BAPyd) libraries

The highly regioselective Buchwald–Hartwig amination at C-2 of the cheap and readily accessible reagent, 2,4-dichloropyridine with a range of anilines and heterocyclic amines is described. This new methodology is robust and provides a facile access to 4-chloro-N-phenylpyridin-2-amines on 0.25 mol scale. These intermediates undergo a further Buchwald–Hartwig amination at higher temperature to enable rapid exploration of the chemical space at C-4 and to provide a library of 2,4-bisaminopyridines.     

Sulfur-directed metal-free and regiospecific methyl C(sp3)–H imidation of thioanisoles

Regiospecific and direct imidation of the methyl C(sp³)–H bond of thioanisoles is realized under mild and metal-free conditions with N-fluorobis(benzenesulfonyl)imide as an oxidant and nitrogen source. Proposed mechanism suggests that thionium ion intermediates and a Pummerer-type reaction are involved. The imidation has advantages such as high step-economy, excellent functionality tolerance, and regiospecificity, giving structurally diverse imidation products.