酪氨酸激酶受体多肽片段在SPR传感芯片表面的磷酸化及其与下游蛋白的相互作用

胰岛素样生长因子-1受体(IGF-1R)信号转导通路与各种正常生理活动及多种疾病发生密切相关.将IGF-1R来源的未磷酸化多肽序列LYASVNPEY固定于表面等离子体共振(SPR)生物传感器专用的CM5芯片表面,以含有相应蛋白激酶的细胞裂解液进行处理,利用SPR生物传感器和抗酪氨酸磷酸化抗体PY20进行酪氨酸磷酸化水平检测.结果表明:PY20能灵敏地检测多肽酪氨酸磷酸化水平变化,即能够区分不同孵育时间以及不同成分的细胞裂解液所引起的多肽酪氨酸磷酸化水平差异;研究了IGF-1R来源多肽的酪氨酸磷酸化对多肽-蛋白间相互作用的影响,证实了只存在磷酸化的多肽与其下游的胰岛素受体底物(IRS-1)相互作用,测定了此磷酸化的多肽与IRS-1的亲和常数(KA)为 (2.02 ± 0.09)×108 L mol-1、结合速率常数(ka)为(2.30±0.15)×106 L mol-1s-1以及解离速率常数(kd)为(1.14±0.13)×10-2 s-1.这种新型的模拟受体的多肽酪氨酸磷酸化研究方法可用于受体磷酸化信号通路的研究以及基于这些信号通路的药物筛选研究.     

基于纳米结构材料的磷酸化蛋白/多肽富集和分析

蛋白质磷酸化是最为广泛的翻译后修饰之一。在生物体液或组织中,许多低丰度的磷酸化蛋白和磷酸化肽是具有高度临床灵敏性和特异性的生物标记物,这些生物分子对许多疾病的检测和病理的阐释可能提供重要的信息。因为蛋白质磷酸化动态可逆且磷酸化蛋白丰度很低,所以很难直接从复杂的生物样品中直接检测到磷酸化蛋白和磷酸化肽。纳米结构材料因其大比表面积、丰富的活性亲合位点和特殊结构,在磷酸化肽和磷酸化蛋白的分离和富集方面已经引起了特别的关注,并成为目前磷酸化蛋白质组学富集和鉴定方面的研究热点。许多介孔、磁性、杂化或化学修饰的亲合材料被研发并用于磷酸化蛋白/多肽的富集与分离;此外,一些多功能纳米结构材料也被研发并用于蛋白质组学中磷酸化蛋白/多肽的快速高效的富集提纯。在这篇综述中,我们专注于纳米结构材料在磷酸化蛋白/多肽富集和提纯方面的最新进展。     

利用电子自旋共振(ESR)技术研究啤酒原辅料自由基及其对麦汁自由基的影响

利用电子自旋共振(ESR)技术研究了啤酒生产原辅料固有自由基含量,发现麦芽自由基量最多,大麦其次,大米中未测到;且自由基存在于皮壳中.全麦麦汁和30%大米辅料麦液都含有羟基自由基和FR1.随糖化进行,羟基自由基上升,FR1下降;但全麦麦汁羟基出现较早,且糖化终了含量较高.     

壳寡糖和磷酸化壳寡糖对Cu(Ⅱ)的络合和缓释性能

以壳寡糖(COS)和磷酸化壳寡糖(PCOS)为原料与铜离子反应,制备了壳寡糖铜(Ⅱ)络合物(COS-Cu)和磷酸化壳寡糖铜(Ⅱ)络合物(PCOS-Cu),讨论了pH、时间、温度和PCOS取代度对络合物吸附量的影响.释放性能表明COS-Cu(Ⅱ)和PCOS-Cu(Ⅱ)均具有缓释性能,且PCOS-Cu(Ⅱ)具有更加均匀的释放速率.     

基于D1蛋白结构的光合作用抑制剂化合物库的组合合成

在所搭建的光合系统Ⅱ (PSⅡ )D1蛋白与先导化合物———氢化脲嘧啶相互作用的复合模型基础上 ,采用固相有机合成的方法平行合成了 8个氢化脲嘧啶类似物 5 ,并用FTIR跟踪了该固相反应 .丙烯酸 (酰氯 )与Wang树脂 1作用得到固载的丙烯酸酯 2 ;2与伯胺经Micheal加成反应生成仲胺 3;随后与异氰酸酯加成得到脲 4;4在酸催化下从树脂上解离的同时关环 ,从而得到氢化脲嘧啶 5 .产物 5的结构经1HNMR ,HPLC和MS所证实 .在此基础上通过变化取代基 ,组合合成了含 9种氢化脲嘧啶的小化合物库 ,它们的结构经GC MS所证实     

光系统Ⅱ反应中心内组氨酸参与给体端次级电子转移反应的人工模拟ESR研究

对高等植物光系统皿(PS)反应中心的超分子模型化合物光诱导电子转移(PET)过程的研究主要集中在原初电子给体P。s。与其还原侧电子受体间的电子转移反应[‘’u．近年来,随着人们对高等植     

介孔氧化铁材料快速、高效富集分离磷酸化肽段和蛋白的新方法研究

以0.1μmol/Lβ-casein磷酸化蛋白酶解液为对象,利用在酸性条件下对PO43-能够特异性吸附的氧化铁材料为新载体,对介孔氧化铁材料富集分离磷酸化肽段的孵育液酸含量、孵育液有机溶剂含量和洗脱液选择的条件进行了优化,结果表明:室温条件下,在含有0.1%乙酸和30%乙腈的孵育液中孵育5min后,经1mol/LNH3.H2O溶液的洗脱,介孔氧化铁可有效地将磷酸化肽段从蛋白酶解液中富集分离。本方法也可以选择性地提取α-casein磷酸化蛋白,实现了简单、快速、高效的磷酸化肽段和蛋白的富集分离。同时,通过MALDI-TOF串级质谱分析,成功地完成了在优化条件下分离出的磷酸化肽段磷酸位点的鉴定。     

半导体光催化模拟光合磷酸化研究(IV)-单色光(红、绿、蓝)对ATP合成的影响

报导了以无机半导体材料(Zn*Cd)S∶Ag-KI为催化剂, 红、绿、蓝3种单色光照下的光照度、照光时间、二磷酸腺苷(ADP) 浓度及催化剂量等对三磷酸腺苷(ATP)合成的影响, 获得了单色光光照度、 ADP 浓度、 Na2HPO4 (Pi)浓度及催化剂量等对ATP合成的最佳条件. 在合适的条件下, ADP 浓度为0.1 mmol/L 时的转化率平均可达到2.6～5.8 %, 绿光在合适的条件下对合成ATP有更高的效率, 说明用无机半导体代替植物叶绿体研究控制光-化学转换是完全可行的.     

固定化金属离子亲和发光二氧化硅纳米粒子的制备及其用于磷酸化蛋白免疫印迹标记

发展了一种能够识别磷酸化蛋白的固定化金属离子亲和发光二氧化硅纳米粒子用于免疫印迹(Western Blot)磷酸化蛋白的标记.首先通过反相微乳液St?ber方法合成了掺杂异硫氰酸荧光素硅烷化衍生物的发光二氧化硅(FITC@SiO2)球形纳米粒子,粒子平均粒径为60 nm.然后通过共聚反应在FITC@SiO2纳米粒子表面...     

LaCl3和PrCl3对菠菜离体叶绿体光合磷酸化作用的影响

LaCl3，PrCl3对离体叶绿体的FeCy光还原活性，随La^3 ，Pr^3 浓度递增而增高，动力学曲线明显呈S形；LaCl3，PrCl3处理叶绿体后，能促进循环( PMS)和非循环( FeCy或MV)光合磷酸化反应，提高叶绿体的偶联程度，增加ADP／O及PC比值，对叶绿体膜上的Mg^2 -ATP酶及偶联因子Ca^2 -ATP酶活力有一定激活作用。这表明LaCl3，PrCl3促进光合作用可能与激活偶联因子活性有关。     

γ射线辐射诱导聚碳硅烷自由基的衰变

利用电子自旋共振波谱(ESR)研究了在N2气中γ射线辐射诱导聚碳硅烷(PCS)自由基的产生和演变行为.ESR谱图分析结果表明,γ射线辐射诱导PCS产生的自由基为硅自由基(≡Si·).低剂量辐照时硅自由基的浓度随吸收剂量的增加而线性增加,硅自由基的辐射化学产额G值约为9,吸收剂量达到200 k Gy后,硅自由基的浓度趋于饱和.室温下硅自由基的浓度随存储时间的延长而逐渐降低,在N2气中存储时硅自由基的半衰期约23 d,在空气中存储时硅自由基的氧化反应导致衰减速率加快,半衰期仅为8 h.温度升高硅自由基衰减速率加快,在N2气中250℃加热处理可以完全清除硅自由基.     

Characterization of x-ray diffraction and electron spin resonance: Effects of sintering time and temperature on bovine hydroxyapatite

The physical and chemical properties of a hydroxyapatite produced by the sintering of bovine bone were investigated by powder x-ray diffraction (PXRD), electron spin resonance (ESR), energy dispersive x-ray spectroscopy (EDX), scanning electron microscopy (SEM), Fourier transform infrared (FTIR), and differential thermal analysis (DTA). A bovine bone powder was sintered at different temperatures ranging from 500 to 1400 °C. The influences of post-irradiation storage on the radiation ESR response of the bovine bone powder before and after sintering were also studied. The results indicate that the sintered bovine bone powder contained hydroxyapatite. Diffraction patterns were sharp and clear based on the (211), (300), and (202) reflections corresponding to bovine hydroxyapatite (BHA), which confirmed the phase purity and high crystalline grade of the BHA produced. The PXRD profile of BHA was dependent on sintering temperatures and times. The molecular formula of BHA was determined by Rietveld analysis showed a similar structure and composition to calcium hydroxyapatite in hexagonal P6₃/m space group a=b=9.435 Å and c=6.895 Å. ESR data showed that the sintering process can decrease the number of free radicals in BHA; it also revealed that the number of free radicals is constant during long storage periods (75 days). The sintering technique described in this study may be used to extract hydroxyapatite from biowaste bovine bone, leading to its application as a bone filler.     

ESR imaging and FTIR study of thermally treated poly(acrylonitrile-butadiene-styrene) (ABS) containing a hindered amine stabilizer: Effect of polymer morphology, and butadiene and stabilizer content

Electron spin resonance imaging (ESRI) was applied to the study of thermal degradation at 393 K of poly(acrylonitrile-butadiene-styrene) (ABS) prepared by emulsion polymerization and containing 25% wt butadiene (ABS-25B). The polymer was doped with 1 or 2% wt Tinuvin 770 as the hindered amine stabilizer (HAS). The spatial distribution of the HAS-derived nitroxide radicals, obtained by 1D ESRI, was initially homogeneous, but became heterogeneous through sample depth with increasing treatment time, t. The spatial variation of ESR line shaping with sample depth was visualized by 2D spectral-spatial ESRI. ESR spectra along the sample depth, obtained by nondestructive (“virtual”) slicing of the 2D images, were used to deduce the relative intensity of nitroxide radicals present in two dynamically distinct sites; the sites were assigned to butadiene-rich (fast component) and SAN-rich domains (slow component), respectively. 1D and 2D ESRI allowed the determination of the extent of degradation within morphologically-distinct domains as a function of sample depth and treatment time. The results from the ESRI experiments were substantiated by attenuated total reflectance (ATR)-FTIR spectroscopy of the outer layer (500 μm thick) of the polymer. Both techniques indicated faster degradation of polymer samples that contained the higher HAS content, 2% wt. Comparison with the results obtained for a parallel study of ABS prepared by mass polymerization and containing 10% wt butadiene (ABS-10B) indicated clearly that the rate of degradation of the polymer prepared by emulsion polymerization (ABS-25B) is significantly reduced. This result can be explained by the formation of cross-linked “composite” networks during emulsion polymerization, which leads to greater thermal stability.     

Direct ESR detection and spin trapping of radicals generated by reaction of oxygen radicals with sulfonated poly(ether ether ketone) (SPEEK) membranes

The stability of membranes under the strong oxidizing conditions in fuel cells is one of the major challenges in the development of fuel cells based on proton exchange membranes (PEMs). This study is centered on the determination of the susceptibility to degradation of SPEEK membranes exposed to OH radicals, using both direct ESR and spin trapping with 5,5-dimethyl-1-pyrroline-1-oxide (DMPO). In order to achieve a complete picture on SPEEK degradation, two types of experiments were performed: 1. UV irradiation at 77 K of SPEEK membranes swollen by aqueous solutions of H₂O₂; 2. UV irradiation of SPEEK membranes swollen by aqueous solutions of H₂O₂ in the presence of DMPO as a spin trap. UV irradiation without oxygen of SPEEK at 77 K in acid or basic form in the presence of H₂O₂/H₂O produced phenoxyl radicals as the predominant radicals detected by direct ESR or spin trapping methods. At pH 4, the oxygen radicals produced phenyl radicals as the predominant species detected by spin trapping methods. The hydroperoxyl radical, as DMPO/OOH adduct, was detected only when the DMPO/OH adduct was absent. The appearance of phenyl and phenoxyl radicals provides the evidence that OH radicals react with the aromatic ring of SPEEK or leading to the scission of its ether bridge.     

四卤化碳: 强碱反应体系顺磁共振研究

单电子转移反应是物理有机化学中十分活跃的研究领域之一,现已发现许多有机反应中都存在着单电子转移过程。探讨不同的单电子转移反应体系,了解其中电子给、受体之间的相互作用,不仅对深入了解反应机理有理论     

Advanced protocol for the detection of irradiated food by electron spin resonance spectroscopy

Using ESR (electron spin resonance) spectroscopy, we found various free radicals in a pepper before and after irradiation. The representative ESR spectrum of the pepper composed of a sextet centered at g=2.0, a singlet at the same g-value and a singlet at g=4.0. This reflects the evidence of three independent radicals in the pepper before irradiation. Upon gamma ray irradiation, a new pair of signals appeared. The progressive saturation behavior (PSB) at various microwave power levels indicates quite different relaxation behaviors of those signals. For the evaluation of radiation-induced radicals and irradiation effects we propose a new protocol using the PSB method. This would call for an advanced protocol for the detection of irradiated foods.     

可见光响应光触媒的测试研究

在液-固相光催化反应体系,采用自旋捕捉-电子自旋共振（ESR）技术,进行可见光响应光触媒的测试研究。在光催化反应体系中加入自由基捕捉剂5-二乙氧基磷酰基-5-甲基-1-吡咯啉-氮氧化物（DEPMP0）,测试不同的光辐射条件下溶液的ESR波谱,测试跟踪光催化反应过程中自由基的动态。本法简便、准确、快捷,可用于光触媒性能的测试以及光催化反应机理的研究。     

ESR study of the molecular orientation and dynamics of stable organic radicals included in the 1‐D organic nanochannels of 2,4,6‐tris‐4‐(chlorophenoxy)‐1,3,5‐triazine

The molecular orientation and dynamics of the organic stable radicals such as 2,2,6,6‐tetramethyl‐1‐piperidinyl‐1‐oxyl (TEMPO) or 4‐hydroxy‐TEMPO (TEMPOL) included in the one‐dimensional (1‐D) organic nanochannels of 2,4,6‐tris‐4‐(chlorophenoxy)‐1,3,5‐triazine (CLPOT) were investigated by examining the inclusion compounds (ICs) diluted by the co‐inclusion of non‐radicals using ESR spectroscopy. Spectral simulation showed that the axial rotation of TEMPO or TEMPOL molecules is excited in the nanochannels with activation energies of 8 and 7 kJ mol^?1, respectively. The rotation axis was estimated to be tilted towards the principal x direction in the axis system of the g ‐tensor of the respective radicals. This is quite different from that for similar ICs in the nanochannels of tris(o‐phenylenedioxy)cyclotriphosphazene (TPP), in which the radicals are axially rotating around the principal axis y of the g‐tensor. The difference is attributed to the larger nanospace of the CLPOT nanochannels. Copyright © 2012 John Wiley & Sons, Ltd.     

BaF2中Vk心的ESR谱分析

电子自旋共振（ＥＳＲ）实验间接表明，γ射线辐照能在室温下在ＢａＦ２微晶中产生Ｆｒｅｎｋｅｌ激子这样的元激发，此元激发受热在１５０℃以下消失。γ射线辐照使ＢａＦ２微晶中Ｆ－离子上的２Ｐ电子跃迁到外层轨道，并与２Ｐ轨道的空穴形成紧束缚的电子－空穴对——Ｆｒｅｎｋｅｌ激子。激子中的电子受热脱离Ｆ原子，这种晶格上的Ｆ０是ＥＳＲ可探测的顺磁色心点缺陷——Ｖｋ心。受γ射线辐照的ＢａＦ２微晶中含有Ｆｒｅｎｋｅｌ激子，受热时Ｆｒｅｎｋｅｌ激子湮灭。Ｖｋ心增多的现象，表现为随温度增高Ｖｋ心的ＥＳＲ信号增强的反常现象。