Subunit‐Specific Labeling of Ubiquitin Chains by Using Sortase: Insights into the Selectivity of Deubiquitinases

Information embedded in different ubiquitin chains is transduced by proteins with ubiquitin‐binding domains (UBDs) and erased by a set of hydrolytic enzymes referred to as deubiquitinases (DUBs). Understanding the selectivity of UBDs and DUBs is necessary for decoding the functions of different ubiquitin chains. Critical to these efforts is the access to chemically defined ubiquitin chains bearing site‐specific fluorescent labels. One approach toward constructing such molecules involves peptide ligation by sortase (SrtA), a bacterial transpeptidase responsible for covalently attaching cell surface proteins to the cell wall. Here, we demonstrate the utility of SrtA in modifying individual subunits of ubiquitin chains. Using ubiquitin derivatives in which an N‐terminal glycine is unveiled after protease‐mediated digestion, we synthesized ubiquitin dimers, trimers, and tetramers with different isopeptide linkages. SrtA was then used in combination with fluorescent depsipeptide substrates to effect the modification of each subunit in a chain. By constructing branched ubiquitin chains with individual subunits tagged with a fluorophore, we provide evidence that the ubiquitin‐specific protease USP15 prefers ubiquitin trimers but has little preference for a particular isopeptide linkage. Our results emphasize the importance of subunit‐specific labeling of ubiquitin chains when studying how DUBs process these chains.     

二代PAMAM树型高分子的荧光现象及其荧光标记应用

首次发现不含荧光团的二代poly(amido amine) (PAMAM) 树型高分子的荧光现象,研究结果表明在无荧光基团的PAMAM树型高分子中,胺类官能团是产生荧光的关键.不同酸度条件下,生成荧光中心的速度不同,而且荧光的稳定性也不相同.同时发现当反复变换酸碱性时PAMAM树型高分子的荧光强度逐渐变弱.由于低代PAMAM树型高分子的良好生物相容性和低廉价格,其有可能成为新型生物荧光标记物.     

多重PCR方法检测食品中转基因成分

根据转基因农作物中最常用的花椰菜花叶病毒启动子 (CaMV 3 5S)和根癌农杆菌终止子(NOS)的序列 ,设计合成了两对不同的引物和相对应的两种荧光双链探针 (FDCP) ,分别建立了多重PCR、应用FDCP的实时荧光PCR同时检测转基因成分 3 5S启动子和NOS终止子的方法 .并利用该套方法对马铃薯、大豆、玉米、甜椒、番茄等实物样品进行了检测 ,发现 1 3份样品中有 6份检出3 5S启动子、NOS终止子 ,其余 7份样品的检测结果为阴性 .表明作者建立的多重PCR方法能有效检测出 3 5S和NOS成分 ,其中多重PCR法具有灵敏度高、特异性好的特点 ,多重荧光PCR法则更为简便、快速、准确     

Histamine determination in tears as an index of safety in use of cosmetic products

To assess ocular irritancy caused by chemical and cosmetic products a reliable method based on evaluation of histamine (Hm) in tears is presented. Hm is measured at picomole levels by HPLC and fluorimetric detection after fluorescamine-HM derivatization. In order to avoid any uncontrolled irritation and stimulation of the conjunctiva during sample collection, a procedure of conjunctiva lavage was developed. A balanced salt solution (50 μl) containing a known amount of Hm-fluorophore as reference standard is instilled in the conjunctiva fornix. After a few seconds 20 μl of tear fluid is collected: 10 μl are immediately analysed and 10 μl after derivatization reaction. In this way it is possible to evaluate tear dilution and to assess Hm content in less than 10 minutes.
In a group of 20 normal subjects Hm has been determined in comparison with that of two volunteers after topical application of 50 μ of 0.2% and 0.4% sodium lauryl sulphate solution. A contact of 30 seconds of the cosmetic ingredient caused an immediate dose-dependent Hm release through a direct cytotoxic damage of cell membranes due to the surfactant action.
Évaluation de l'irritation occulaire due aux cosmétiques par la détermination de l'histamine lacrimale     

Miscibility characterization of SMA/SAN and SMA/PMMA blends by differential scanning calorimetry and fluorescence techniques

In this study, styrene‐maleic anhydride (SMA) copolymer was modified by monoesterification method with 9‐(hydroxymethyl)anthracene fluorophore to prepare a fluorescent anthracene labeled SMA (SMA‐An) material. The latter was then characterized by attenuated total reflection (ATR) and thermogravimetric analysis (TGA) techniques. In the second step of this work, SMA‐An was added to SMA/[Styrene‐Acrylonitrile Copolymer (SAN)] and SMA/[Poly(methyl methacrylate) (PMMA)] blends to investigate the miscibility of these blends at the molecular level. The miscibility of SMA/PMMA blends was characterized using fluorescence quenching of anthracene by the succinic anhydride and succinic acid functions on SMA macromolecule itself. © 2007 Wiley Periodicals, Inc. J Appl Polym Sci 2007     

可见波段Ar+激光诱导血红蛋白荧光光谱特性研究

用可见波段、不同波长的Ar^ 激光激发同一浓度血红蛋白溶液产生荧光光谱的研究结果表明，血红蛋白在628nm附近存在一个较强的荧光谱峰，且随激励激光波长的红移其相对光强依次增大，从而解释了临床上用He-Ne激光治疗疗效显著的物理机理。理论研究结果认为该血红蛋白的荧光光谱主要是血红蛋白中存在的卟啉类荧光团的贡献。还利用475．6nm的激光分别激发浓度为1％～7％的血红蛋白溶液，根据所获得的荧光光谱发现，其谱峰位置几乎不随样品浓度的改变而改变，该结果显示了激光与普通光对生物大分子存在明显不同的作用特性。     

含全氟丙基的四取代嘧啶的制备及光学性质研究

利用“一锅多组分”方法合成了一种含全氟丙基的四取代嘧啶(DMA-PM),并用溶液挥发法培养出两种晶体材料(A和B),该晶体材料作为一种迷你荧光体,分别具有明显的蓝色CIE(0.19,0.12)和黄色CIE(0.49,0.51)荧光,它们的最大发射波长分别为435 nm和563 nm．晶体学研究表明,这种有趣的发射现象是源于它们晶体结构中,依赖于弱相互作用的发射和构象平面化．     

免疫层析试纸条标记探针研究进展

免疫层析试纸条(Immunochromatographic test strips,ICTSs)有效结合了层析法卓越的分离能力和常规免疫分析技术的高度特异性,加之其便携性,该技术无疑为灵敏、定量现场检测提供了一个理想的平台。近年来随着检测方法和技术的不断革新,包括生色、发光及电化学信号产生型等在内的多种探针不断涌现,它们或直接共价结合靶标蛋白或装载于纳米颗粒中用于免疫层析试纸条的标记。本文旨在总结归纳当前ICTSs标记探针的最新进展,详尽解析各类探针性质、发展、优略势及其在生物分析中的应用,以期为研究者在设计试纸条时适宜探针的选择提供有力的技术支持。     

Highly Fluorescent Thienoviologen‐Based Polymer Gels for Single Layer Electrofluorochromic Devices

A highly fluorescent electrofluorochromic gel with quantum yields as high as 67% is prepared by incorporating the thienoviologen fluorophore 4,4′‐(2,2′‐bithiophene‐5,5′‐diyl)bis(1‐nonylpridinium)bistriflimide into a polymeric matrix. The fluorescent emission spectrum of the gel at low percentages of thienoviologen shows a strong band at 530 nm. A new intense fluorescence band at 630 nm can be induced by fluorophore aggregation. Single layer electrofluorochromic devices were readily prepared by sandwiching the polymer gels between two indium tin oxide (ITO) electrodes. The fluorescence intensity can be easily modulated between a fluorescent and a quenched state, in a wide visible spectral range, by direct electrochemical reduction of the thienoviologen fluorophore. It exhibits three reduction states, each with different emission properties. The reversible interconversion among these states leads to a high electrofluorochromic stability of the device, exhibiting switching times of a few seconds and, to the best of our knowledge, the highest contrast ratio (337).