Arabidopsis 2010: Development of an Arabidopsis Proteome Chip

由于所研究的大多数蛋白质缺乏可辨别的酶或生化功能，蛋白质的大规模表征将取决于与它们相关的分子的鉴定。蛋白质与其他蛋白质、小分子、脂类或核酸的结合对其定位、活性、稳定性和细胞功能有深远的影响。为了在全球范围内阐明拟南芥蛋白质组的生化活性，该项目将产生一个包含10,000个拟南芥ORF的拟南芥表达集合(ATEC)，并将其融合到串联亲和纯化(TAP)标签中。将使用优化的植物表达系统来生产和纯化这些蛋白质。这些蛋白质将被印刷在不同的表面以产生拟南芥蛋白质芯片，然后将被筛选用于各种生化分析，包括蛋白质-蛋白质相互作用，蛋白质-磷脂相互作用，蛋白质-核酸相互作用，蛋白质-小分子相互作用，以及鉴定激酶和其他酶的底物。产生的试剂和方案将通过项目网站(http://plants.gersteinlab.org/)，、拟南芥生物资源中心和拟南芥信息资源)提供给科学界。广泛影响：大规模生物学领域的快速进展为大规模阐明生物功能以及构建和理解生物网络提供了机会。由于基因的生化功能是通过其编码的蛋白质来体现的，因此蛋白质组学的新工具的开发受到了极大的重视。蛋白质微阵列为同时分析数千个单独的蛋白质样本提供了一个强大而方便的平台，便于在高度特定的条件下设计复杂和可重复性的生化实验。因此，本项目所做的努力最终将为旨在高通量研究拟南芥蛋白质功能和阐明复杂生物网络的各种应用提供有价值的资源。了解这些网络以及它们是如何控制的，可能有助于将其应用于胁迫、响应、能量生产等。此外，本研究开发的方法和获得的信息也可以直接应用于其他农业和园艺重要植物的分析。最后，该项目将整合对学生和博士后在蛋白质组学前沿领域的研究和教育培训，并将提供一个独特的机会，提高少数族裔本科生、公立高中学生和教师对植物蛋白质组学研究的认识和重要性。

Since the majority of proteins studied lack discernable enzymatic or biochemical functions, large-scale characterization of proteins will depend on the identification of molecules that associate with them. The association of a protein with other proteins, small molecules, lipids, or nucleic acids has profound effects on its localization, activity, stability, and cellular function. To elucidate the biochemical activities of the Arabidopsis proteome on a global scale, the project will generate an Arabidopsis thaliana expression collection (ATEC) containing 10,000 Arabidopsis ORFs, fused to tandem affinity purification (TAP) tag. An optimized plant-based expression system will be used to produce and purify these proteins. The proteins will be printed on various surfaces to produce Arabidopsis protein chips which will then be screened for various biochemical assays, including protein-protein interactions, protein-phospholipids interactions, protein-nucleic acid interactions, protein-small molecule interactions, and the identification of substrates for kinases and other enzymes. The reagents and protocols generated will be made available to the scientific community through the project website (http://plants.gersteinlab.org/), the Arabidopsis Biological Resource Center (ABRC) and the Arabidopsis Information Resource (TAIR).Broader Impacts: The rapid progress in the field of large-scale biology has provided the opportunity to elucidate biological function on a large scale and to construct and understand biological networks. Since the biochemical function of a gene is manifested through its encoded protein, much emphasis has been placed on the development of new tools for proteomics. A protein microarray provides a robust and convenient platform for the simultaneous analysis of thousands of individual protein samples, facilitating the design of sophisticated and reproducible biochemical experiments under highly specific conditions. Therefore, the efforts undertaken in this project will ultimately provide a valuable resource for a variety of applications aimed at the high-throughput study of protein function in Arabidopsis and for the elucidation of complex biological networks. Understanding these networks and how they are controlled is likely to be enabling for application to stress, response, energy production etc. In addition, the methods developed and information gained from this study can also be directly applied to the analysis of other agriculturally and horticulturally important plants. Finally, the project will integrate research and educational training for students and post-doctoral associates in the cutting-edge field of proteomics and will provide a unique opportunity to elevate the awareness and importance of plant proteomics research to minority undergraduate students, public high school students and teachers.