串联质谱谱库搜索鉴定技术综述

介绍蛋白质质谱鉴定领域的串联质谱谱库搜索技术,分析谱库搜索技术的基本原理,从多个角度阐述当前主流的谱库搜索方法,并对其优缺点进行比较,针对目前蛋白质质谱鉴定技术所存在的优缺点,设计其改进思路,论述目前谱库搜索技术所面临的问题以及未来的研究趋势。     

酵母蛋白网络的功能模块与关键蛋白研究

蛋白交互网络在各种细胞功能和生命过程中发挥着至关重要的作用。对它的结构特征进行分析吸引了众多科研人员的关注,功能模块与关键蛋白的识别是其中的重要研究主题。本文主要使用模拟退火算法分析了酵母蛋白交互网络的功能模块,同时结合使用多种中心化指标识别了其中的关键蛋白,并讨论了这些功能模块和关键蛋白的生物学意义。     

Identification and Molecular Cloning of Putative Odorant-Binding Proteins from the American Palm Weevil, Rhynchophorus palmarum L.

We have identified and cloned the cDNAs encoding two odorant-binding proteins (OBPs) from the American palm weevil (APW) Rhynchophorus palmarum (Coleoptera, Curculionidae). Degenerate primers were designed from the N-terminal sequences and were used in polymerase chain reaction (PCR) in order to obtain full-length sequences in both males and females. In both sexes, two different cDNAs were obtained, encoding 123 and 115 amino acid-deduced sequences. Each sequence showed few amino acid differences between the sexes. The proteins were named RpalOBP2 and RpalOBP4 for male, RpalOBP2' and RpalOBP4' for female, with the types 2 and 4 presenting only 34% identities. These proteins shared high identity with previously described coleopteran OBPs. In native gels, RpalOBP2 clearly separated into two bands and RpalOBP4 into three bands, suggesting the presence of several conformational isomers. Thus, OBP diversity in this species may rely on both the presence of OBPs from different classes and the occurrence of isoforms for each OBP.     

Characterization of Fractionated Soy Proteins Produced by a New Simplified Procedure

It was possible to fractionate soy protein into two soy protein isolate fractions (>90% protein) enriched in either glycinin or β-conglycinin by using a new simplified procedure (referred to as the Deak procedure) employing CaCl₂ and NaHSO₃. The Deak procedure produced fractions with higher yields of solids, protein, and isoflavones, and similar protein purities as well as improved functional properties compared to fractions recovered by established, more complex soy protein fractionation procedures. The Deak glycinin-rich fraction comprised 15.5% of the solids, 24.4% of the protein, and 20.5% of the isoflavones in the starting soy flour, whereas the glycinin-rich fraction of the established procedure (Wu procedure) comprised only 11.6% of the solids, 22.3% of the protein, and 9.6% of the isoflavones. The Deak β-conglycinin-rich fraction comprised 23.1% of the solids, 37.1% of the protein, and 37.5% of the isoflavones in the starting soy flour, whereas the Wu β-conglycinin-rich fraction comprised only 11.5% of the solids, 18.5% of the protein, and 3.3% of the isoflavones. Protein purities were >80% for both fractions when using both procedures. The Wu procedure produced protein fractions with slightly higher solubilities and similar surface hydrophobicities; whereas, the fractions produced by the Deak procedure had superior emulsification and foaming properties and similar dynamic viscosity behaviors.     

DDI-microFIA--A readily configurable microarray-fluorescence immunoassay based on DNA-directed immobilization of proteins

We describe a chip-based immunoassay for multiplex antigen detection, based on the self-assembly of semi-synthetic DNA-protein conjugates to generate an easily configurable protein microarray. The general principle of this microarray-fluorescence immunoassay (microFIA) is similar to that of a two-sided (sandwich) immunoassay. However, covalent single-stranded DNA-streptavidin conjugates are employed for the efficient immobilization of biotinylated capture antibodies through hybridization to complementary surface-bound DNA oligomers. In a model system, we use the DNA-directed immobilization (DDI) of antibodies to generate an antibody microarray for the parallel detection of the tumor marker human carcinoembryonic antigen (CEA), recombinant mistletoe lectin rViscumin (rVis), ceruloplasmin (CEP), and complement-1-inactivator (C1A) in human blood serum samples. Detection limits down to 400 pg mL(-1) are reached. In addition, we describe a method for the internal standardization of protein microarray analyses, based on the simultaneous measurement of constant amounts of the blood proteins CEP and C1 A, intrinsically present in human serum, to compensate for interexperimental variations usually occurring in microarray analyses. The standardization leads to a significantly higher data reliability and reproducibility in intra- and interassay measurements. We further demonstrate that the DDI-microFIA can also be carried out in a single step by tagging of the analyte simultaneously with both capture and detection antibody and subsequent immobilization of the immunocomplex formed, on the DNA microarray capture matrix. This protocol significantly reduces handling time and costs of analysis.     

蛋白质双水相液液平衡的热力学模型与研究进展

双水相体系由于性质温和、操作方便、分离效率高,以及不会导致被分离物质的破坏和失活等优点,被广泛应用于生物大分子物质的分离和纯化.目前主要应用的双水相体系有高聚物-高聚物体系、高聚物-电解质体系和近年来刚发展起来的胶束双水相体系.针对此3种不同类型的分离体系,分析了蛋白质在其中分配系数的热力学模型,希望对进一步研究有所帮助和启发.     

大豆蛋白酶解产物降胆固醇活性的初步研究

研究了用AS1.398和Alcalase两种蛋白酶制备的水解度为10%～24%的大豆蛋白酶解产物的降胆固醇活性.结果表明:用Alcalase酶水解大豆蛋白,水解度为18%的产物降胆固醇活性最高,添加量为5 mg/mL时,对胆固醇胶束溶解度的抑制率为48.39%,动物实验表明该产品在灌喂剂量大于1 g/(kg·d)时,对喂饲高脂饲料小鼠的血清总胆固醇降低作用比较显著(P＜0.025),30 d后降低幅度可达30%以上.     

基于Z曲线的SARS冠状病毒特征的研究

刺突（Spike）蛋白是SARS（Severe Acute Respiratory Syndmme）冠状病毒表面最重要的膜蛋白，它通过与被感染细胞的受体结合来作为感染的媒介。受此启发，该文利用Z曲线方法，对S蛋白及SARS受体序列进行探究，捕捉到一种SARS病毒的可视化特征。大量序列比对的结果证明该特征是SARS病毒所特有的。将这种特征图谱应用到SARS病毒的检测中，可以提供一种非标准的、简单、直观的检验方法，能够诊断一些利用标准方法难以判断的SARS病例。由此可见，Z曲线作为一种基因序列的几何学研究途径是一种有效的研究方法。     

水解松仁蛋白用酶的最佳工艺条件

为了充分利用松仁中的蛋白质，将其酶解制备活性多肽，分别对7种蛋白酶进行水解实验。用pH-stat法控制水解度，用福林-酚法测定吸光度，计算氮溶指数。通过正交实验得出如下结果：最佳用酶是Asl．398中性蛋白酶。最佳水解条件为：温度50℃，pH=7．0，底物浓度1．2％，酶与底物比4000U／g蛋白。     

超微粉碎小麦的蛋白质变化探讨

采用自行设计制造的CH—300型机械冲击式粉碎机对小麦进行粉碎加工、分级，通过工艺参数调整，选出两种加工分级后的产品进行分析，结果表明：加工后的小麦平均粒径可达5．85μm，形貌呈球对称状；其蛋白质含量随粒径变细而增加，当D9小于10μm时，蛋白质含量骤然增大一倍多，从而为小麦的特殊使用要求提供了深加工的途径。