EAGER: Tool development for proteomics and environmental metaproteomics

EAGER：蛋白质组学和环境宏蛋白质组学工具开发

• 批准号: 0938190
• 负责人: Brian Palenik
• 金额: $ 8.76万
• 依托单位: University of California-San Diego Scripps Inst of Oceanography
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-08-01 至 2011-07-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=0938190&HistoricalAwards=false
• 关键词: EAGER; Tool; development; proteomics; environmental

This EAGER project to Brian Palenik, UC San Diego, is for tools development to identify previously unknown, putative genes in metagenomic sequence data. Metagenomic sequences represent the full complement of microbial genomes in a community and are difficult to analyse because of their complexity. Even the genomes of cultured organisms are difficult to annotate as typically 50% of the genome sequence can contain unknown or novel genes. Proteomics uses mass spectrometry to analyze peptides derived from the proteolysis of samples. This analysis allows for the determination of the presence or absence of specific proteins in the original sample. In some forms, proteomics can use genomic databases as a starting point for peptide and protein identification. More recently, a form of "reverse proteomics" is being tested where mass spectrometry data are used to improve whole genome annotation by demonstrating the presence of proteins not predicted in the initial genome annotation. This technology is now beginning to show promise in the analysis of complex environmental samples, such as metaproteomic samples. This type of community-level analysis can lead to new information about not only the biodiversity but the functioning of ecosystems. However, the coupling of mass spectrometry and genome and metagenome data to analyze environmental samples still requires tool development. This development is best done when the metagenome and metaproteome sampling can be coordinated to optimize and test the tools. This PI is developing several tools for analyzing metaproteomes using metagenomics data obtained from the same site. The tools include computational procedures to discover new open reading frames and genes, methods to identify which species are present without DNA sequencing and an approach for analysing post-translational modifications in Synechococcus metagenomes. This aspect of the work is higher risk than the other aims but is important because this will allow them to evaluate which genes in the metagenome are active. This project represents a novel interdisciplinary collaboration between a marine microbiologist and a mass spectroscopist, and it is timely as it takes advantage of ongoing metagenome sequencing funded by another NSF proposal. If successful, these tools will allow for the identification of activated genes within communities. Further, new or novel genes will be identified in the metagenomes and the products of these novel genes will be identified. This research team will focus on marine cyanobacterial communities as the test metagenome. The PI has already sequenced the genomes of two Synechococcus (marine cyanobacteria) and he will use these as reference genomes for the metagenomic/metaproteomic analyses. Synechococcus assemblages in the ocean are not very diverse, therefore Synechococcus metagenomes are some of the best metagenomes for these tests because these populations can be enriched from seawater to produce a nearly pure samples for proteomic analysis. The PI is overseeing training of two post-docs as a result of this award.

这个急切的向圣地亚哥加州大学布莱恩·帕雷尼克（Brian Palenik）的项目旨在识别宏基因组序列数据中以前未知的假定基因。宏基因组序列代表了一个社区中微生物基因组的完整补体，并且由于其复杂性而难以分析。即使是培养生物的基因组也很难注释，因为通常50％的基因组序列可能包含未知或新颖的基因。蛋白质组学使用质谱法来分析源自样品蛋白水解的肽。该分析允许确定原始样品中特定蛋白质的存在或不存在。在某些形式中，蛋白质组学可以使用基因组数据库作为肽和蛋白质鉴定的起点。最近，正在测试一种“反蛋白质组学”的一种形式，其中使用质谱数据来通过证明在初始基因组注释中未预测的蛋白质的存在来改善整个基因组注释。现在，这项技术开始在分析复杂环境样本（例如元蛋白质组学样本）的分析中表现出希望。这种类型的社区级别分析不仅会导致有关生物多样性的新信息，而且还会导致生态系统的功能。但是，质谱和基因组和元基因组数据的耦合仍需要工具开发。当可以协调元基因组和荟萃蛋白组采样以优化和测试工具时，最好完成此开发。该PI使用从同一站点获得的宏基因组学数据开发了几种用于分析元蛋白质组的工具。这些工具包括发现新的开放式阅读框和基因的计算程序，识别没有DNA测序的物种的方法，以及一种分析Synechococcus元基因组中翻译后修饰的方法。工作的这一方面比其他目标更高，但很重要，因为这将使他们能够评估元基因组中的哪些基因活跃。该项目代表了海洋微生物学家和质谱学家之间的一种新颖的跨学科合作，并且它及时，因为它利用了由另一项NSF提案资助的持续元素测序。如果成功，这些工具将允许在社区内识别激活的基因。此外，将在宏基因组中鉴定出新的或新颖的基因，并且将鉴定出这些新基因的产物。该研究团队将专注于海洋蓝细菌群落作为测试元基因组。 PI已经测序了两个链神经（海洋蓝细菌）的基因组，他将使用这些基因组作为元基因组/元蛋白质组分析的参考基因组。 海洋中的综合菌组合不是很多样化，因此元基因组是这些测试的一些最佳元基因组之一，因为这些种群可以从海水中富集，以产生几乎纯净的样品进行蛋白质组学分析。由于该奖项，PI正在监督两个后训练的培训。