酪氨酸磷酸化蛋白质组学技术研究进展及其在生物医学研究中的应用

在酪氨酸磷酸化蛋白质组学的研究过程中,酪氨酸磷酸化位点的富集是最重要的一步。目前常用的富集方法是抗体亲和富集或SH2 superbinder富集。此外,通过质谱与生物信息学等技术,可实现大规模酪氨酸磷酸化位点的鉴定。对酪氨酸磷酸化蛋白质组学进行深度覆盖研究,揭示癌症发生发展过程中失调的激酶,将有助于深入理解癌症的发生发展过程;且由于75%的致癌基因是酪氨酸激酶基因,酪氨酸激酶抑制剂作为抗癌药物受到了越来越多的关注。应用酪氨酸磷酸化蛋白质组学技术,可以鉴定与癌症等重大疾病相关的酪氨酸激酶,从而帮助找到酪氨酸激酶抑制剂。总之,酪氨酸磷酸化蛋白质组学技术可以在酪氨酸激酶鉴定、酪氨酸激酶抑制剂研究及酪氨酸磷酸化信号通路研究等生物医学领域中得到很好的应用。     

磷酸化蛋白质组学研究中磷酸化肽段富集与分离方法研究进展

对磷酸化蛋白质组（phosphoproteome）进行系统深入的研究依赖于高重复性和特异性的磷酸化肽段富集与分离方法。目前发展了多种不同原理的磷酸化肽段富集方法,它们往往具有不同的选择性和特异性,因此,根据不同的研究目的选择最适合的富集方法显得尤为重要。本文综述了基于亲和色谱法（affinity chromatography）、免疫沉淀法（immunoprecipitation）、化学衍生法（chemical derivatization）、色谱法（chromatography）和其他新发展方法的磷酸化肽段富集方法,详细介绍了各自的优缺点及相关的优化与改进策略。此外,还简单介绍了磷酸化肽段富集与预分方法的不同组合的研究进展。     

磷酸化蛋白质组研究中的富集策略

蛋白质的磷酸化与去磷酸化过程,调控着包括信号转换、基因表达、细胞周期等诸多细胞过程。因此,对蛋白质磷酸化修饰的分析是蛋白质组研究中的重要内容。但由于磷酸化蛋白的丰度较低,难以用质谱直接检测。为了解决这个问题,改善质谱对磷酸肽的信号响应,需要对磷酸化蛋白质或磷酸肽进行富集。目前主要的富集方法包括免疫沉淀、固相金属离子亲和色谱、金属氧化物/氢氧化物亲和色谱等。     

Phos-tag技术在磷酸化蛋白质组学中的应用

Phos-tag是新研制出的一种对磷酸基团具有特殊亲和力的化合物。由于其对磷酸化蛋白质具有高特异性、高亲和力等特点使其迅速在磷酸化蛋白质的检测、分离和纯化等方面得到广泛的应用。本文综述了Phos-tag的化学性质、原理及其近年来在磷酸化蛋白质组学中的应用,并与传统的磷酸化蛋白质组学研究技术做了比较,对未来磷酸化蛋白质组学的研究技术作了展望。     

金属氧化物在磷酸化蛋白质组学研究中的应用

磷酸化肽段的高效富集是磷酸化蛋白质组学中的重要任务,通过综述纳米金属氧化物在磷酸化蛋白组学中的应用,了解磷酸化蛋白组学研究中常用的磷酸化肽段富集材料和方法.该文介绍纳米金属氧化物、核壳结构纳米磁性材料以及金属离子在磷酸化蛋白组学中的富集效果.纳米金属氧化物材料由于其特殊理化性质在富集磷酸化肽段研究中具有特异性强、选择性好特点,在磷酸化蛋白组学研究中得到了广泛应用.虽然纳米金属氧化物在磷酸化蛋白组学的研究中仍然处于起步阶段,可重复性和全面富集仍面临挑战,但是在未来的亚细胞蛋白组学和功能蛋白组学研究中,纳米金属氧化物及其衍生物仍然会起着重要作用.     

中国蛋白质组学研究进展——以人类肝脏蛋白质组计划和蛋白质组学技术发展为主题

蛋白质组学是对细胞或生物体全部蛋白质的系统鉴定、定量并阐释其生物学功能的学科.自21世纪初期开始,随着高精度、高灵敏度和快速扫描质谱仪的出现和快速发展以及微量蛋白质组样品高效分离技术的进步,蛋白质组学获得了快速发展,并在生理过程与病理机制研究等几乎所有生命科学研究领域得到了广泛的应用.过去10年,中国蛋白质组学研究在政府的支持和广大蛋白质组学研究人员的努力下呈现出腾飞式的发展态势.本文综述了人类肝脏蛋白质组计划和2010~2013年中国蛋白质组学技术的发展.     

植物蛋白质磷酸化的研究技术

本文介绍植物蛋白质磷酸化研究的技术及其应用情况,并对这些技术的应用前景作了展望。     

磷酸化蛋白质组学分析和定量技术的研究进展

蛋白质的磷酸化是一种可逆性的蛋白质翻译后修饰,在生物体内起着极为重要的作用.近年来蛋白质翻译后修饰日益成为蛋白质组研究的热点之一.定量磷酸化蛋白质组学方法和技术的快速发展为研究蛋白质磷酸化时空动态变化,更好地了解生物学功能调节网络奠定了坚实的基础.作为蛋白质组学研究的一个重要组成部分,定量磷酸化蛋白质组学因其磷酸化蛋白质所具有的独特特征,在技术和方法研究方面将面临更为严峻的挑战.综述了磷酸化蛋白质组学定量的一些分析技术和方法的发展现状、优缺点以及未来的发展趋势.     

生物质谱及其在蛋白质组学研究中的应用

生物质谱是蛋白质组学研究必不可少的关键技术。近年来,生物质谱在鉴定通量、分辨率和灵敏度等方面均有质的飞跃,从而促进了蛋白质组研究各个领域的飞速发展。本文就生物质谱技术的原理、技术和仪器发展现状,及其在蛋白质组学研究中的应用进展作一简要的综述。     

蛋白质组学及其在植物科学研究中的应用

蛋白质组学是后基因组时代出现的一个新兴研究领域。简要介绍了蛋白质组学的研究意义及其研究内容与技术手段,及蛋白质组研究在植物科学中的应用。     

Proteomic and phosphoproteomic analyses of NaCl stress-responsive proteins in Arabidopsis roots

Salt is one of the major abiotic stresses limiting the productivity and the geographical distribution of crops. To gain a better understanding of NaCl stress responses in model plant Arabidopsis roots, the protein changes in the abundance (Coomassie Brilliant Blue R-350 stain) and phosphorylation (Pro-Q Diamond stain) were examined using two-dimensional electrophoresis coupled with mass spectrometry (MS). Seventeen unique proteins differentially changed in abundance, phosphorylation, or both in response to NaCl. Nonsynchronous differences were found between total proteins and phosphorylated proteins. Protein synthesis, proteolysis, post-translational modifications, and isoforms might cause the differential protein redundancies. The identified proteins are involved in binding, catalysis, signal transduction, transport, metabolisms of cell wall and energy, and reactive oxygen species (ROS) scavenging and defense. These protein changes provide new avenues of investigation into the underlying salt stress response in Arabidopsis roots and demonstrate the advantages of proteomic approach in plant biology studies.     

Structure of Carbohydrate Moiety in Asterosaponin A. Isolation of Three New Disaccharides

ABSTRACT

This study aimed to investigate the role of serine/threonine kinase PkaE in Streptomyces coelicolor A3(2). Liquid chromatography tandem mass spectrometry was performed for comparative phosphoproteome and proteome analyses of S. coelicolor A3(2), followed by an in vitro phosphorylation assay. Actinorhodin production in the pkaE deletion mutant was lower than that in wild-type S. coelicolor A3(2), and the spores of the pkaE deletion mutant were damaged. Furthermore, phosphoproteome analysis revealed that 6 proteins were significantly differentially hypophosphorylated in pkaE deletion mutant (p < 0.05, fold-change ≤ 0.66), including BldG and FtsZ. In addition, the in vitro phosphorylation assay revealed that PkaE phosphorylated FtsZ. Comparative proteome analysis revealed 362 differentially expressed proteins (p < 0.05) and six downregulated proteins in the pkaE deletion mutant involved in actinorhodin biosynthesis. Gene ontology enrichment analysis revealed that PkaE participates in various biological and cellular processes. Hence, S. coelicolor PkaE participates in actinorhodin biosynthesis and morphogenesis.     

Identification of Two Protein-Signaling States Delineating Transcriptionally Heterogeneous Human Medulloblastoma

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New approaches towards integrated proteomic databases and depositories

Since the publication of the human genome, two key points have emerged. First, it is still not certain which regions of the genome code for proteins. Second, the number of discrete protein-coding genes is far fewer than the number of different proteins. Proteomics has the potential to address some of these postgenomic issues if the obstacles that we face can be overcome in our efforts to combine proteomic and genomic data. There are many challenges associated with high-throughput and high-output proteomic technologies. Consequently, for proteomics to continue at its current growth rate, new approaches must be developed to ease data management and data mining. Initiatives have been launched to develop standard data formats for exchanging mass spectrometry proteomic data, including the Proteomics Standards Initiative formed by the Human Proteome Organization. Databases such as SwissProt and Uniprot are publicly available repositories for protein sequences annotated for function, subcellular location and known potential post-translational modifications. The availability of bioinformatics solutions is crucial for proteomics technologies to fulfil their promise of adding further definition to the functional output of the human genome. The aim of the Oxford Genome Anatomy Project is to provide a framework for integrating molecular, cellular, phenotypic and clinical information with experimental genetic and proteomics data. This perspective also discusses models to make the Oxford Genome Anatomy Project accessible and beneficial for academic and commercial research and development.     

Quantitative analysis of representative proteome components and clustering of Helicobacter pylori clinical strains

BACKGROUND: Several Helicobacter pylori proteins have been reported to be associated with severe symptoms of gastric disease. However, expression levels of most of these disease-associated proteins require further evaluation in order to clarify their relationships with gastric disease patterns. Representative proteome components of 71 clinical isolates of H. pylori were analyzed quantitatively to determine whether the protein expression levels were associated with gastric diseases and to cluster clinical isolates. METHODS: After two-dimensional electrophoresis (2-DE) of H. pylori isolates, spot intensities were analyzed using pdquest 2-D Gel Analysis Software. The intensities of 10 representative protein spots, identified by peptide fingerprinting using matrix assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF-MS) or peptide sequencing using quadrupole TOF MS, were subjected to the nonparametric Mann-Whitney test and hierarchical agglomerative cluster analysis. The relationship between clusters and gastric diseases was analyzed by the chi-squared test. RESULTS: Although the spot intensities of the 10 representative proteins were highly variable within each gastric disease group, the expression levels of CagA, UreB, GroEL, EF-Tu, EF-P, TagD, and FldA showed some significant differences among the gastric disease patterns. On the basis of the 10 target protein intensities, hierarchical agglomerative cluster analysis generated a dendrogram with clusters indicative of chronic gastritis/gastric cancers and gastric/duodenal ulcers. CONCLUSION: These results indicated that quantitative analysis of proteome components is a feasible method for examining disease-associated proteins and clustering clinical strains of H. pylori.