Arabidopsis 2010: Novel Folate-Related Proteins Shared by Plants and Prokaryotes

拟南芥 2010：植物和原核生物共有的新型叶酸相关蛋白

• 批准号: 0839926
• 负责人: Andrew Hanson
• 金额: $ 50.72万
• 依托单位: University of Florida
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-05-01 至 2013-04-30
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=0839926&HistoricalAwards=false
• 关键词: Arabidopsis; 2010; Novel; Folate; Related

Scientific Merit:Genome sequencing is transforming discovery in metabolic biochemistry by enabling comparative genomics (phylogenomics) and unprecedented information flow between plant and prokaryote research. This Arabidopsis 2010 project uses the power of extensive biological databases to discover the function of new genes involved in folate metabolism. Folates are small, organic molecules that act as essential cofactors in conjunction with proteins to enable enzymatic reactions that are critical to all forms of life. Folates are involved in reactions that transfer one carbon (C1) units at different levels of oxidation. Despite decades of classical research on folates as cofactors in C1 metabolism, phylogenomics reveals unknown proteins in all kingdoms of life that are candidates for novel folate-dependent functions. This phylogenomics-driven project aims to establish the functions of two such novel proteins in plants and bacteria (YgfZ and APE157), to clarify that of a third (TrmE), and to advance use of phylogenomics in the metabolic biochemistry community. YgfZ proteins are often essential to viability and appear to have a role in iron-sulfur (Fe/S) cluster assembly or repair. A working hypothesis is that YgfZ removes C1 moieties blocking Fe/S cluster binding sites. This hypothesis will be tested by mutational, biochemical, and biophysical strategies in bacteria and Arabidopsis. Genetic and comparative biochemical evidence suggests that TrmE proteins facilitate addition of C1 blocking groups to Fe/S proteins. Mutational and biochemical approaches will be used to test this idea. Structural and genomic context information indicates that APE157 proteins functionally replace the crucial folate recycling enzyme 5-formyltetrahydrofolate cycloligase (5-FCL) by mediating a different reaction. For both prokaryotic and plant APE157 proteins, this hypothesis will be tested by combining genetics, metabolite profiling, isotope labeling, and enzymology. Characterizing YgfZ as a ubiquitous folate-dependent protein involved in Fe/S cluster metabolism would be a new departure in folate biochemistry, as would establishing a folate connection between TrmE, YgfZ, and Fe/S enzymes. Identifying APE157 as a widespread alternative to 5-FCL is an unexplored aspect in folate biochemistry of general importance. Broader Impact:Although phylogenomics is a powerful and cost-effective way to discover unknown metabolic genes in prokaryotes and plants it is currently underused. To promote the application of phylogenomics in metabolic biochemistry, this project will develop a two-day workshop using the SEED database and its tools. This workshop will be presented at two major universities per year for the duration of the award to a target group of plant and prokaryotic biochemists, at PhD student, postdoctoral, and faculty levels. In addition to this wider outreach, this project will contribute to the training of at least two PhD students and two postdoctorals in the laboratories of the PI and Co-PI, and provide undergraduate research opportunities.Data and information generated through this project will be made available through the Arabidopsis Information Resource (TAIR: www.arabidopsis.org). Seed stocks and any unique DNA materials will be made available through ABRC. http://www.biosci.ohio-state.edu/~plantbio/Facilities/abrc/abrchome.htm

科学价值：基因组测序正在通过实现比较基因组学（基因组学）和植物与原核生物研究之间前所未有的信息流来改变代谢生物化学的发现。这个拟南芥2010项目利用广泛的生物数据库的力量来发现参与叶酸代谢的新基因的功能。叶酸是一种小的有机分子，它与蛋白质一起作为重要的辅助因子，使酶反应成为所有生命形式的关键。叶酸参与了以不同氧化水平转移一个碳（C1）单位的反应。尽管数十年来对叶酸作为C1代谢中的辅助因子的经典研究，但基因组学揭示了所有生命王国中的未知蛋白质，这些蛋白质是新的叶酸依赖性功能的候选者。该项目旨在确定植物和细菌中两种新蛋白质（YgfZ和APE 157）的功能，阐明第三种蛋白质（TrmE）的功能，并推动代谢生物化学领域中的代谢基因组学的应用。YgfZ蛋白通常对生存力至关重要，并且似乎在铁-硫（Fe/S）簇组装或修复中起作用。一个工作假设是，YgfZ去除C1部分阻断Fe/S簇结合位点。这一假设将在细菌和拟南芥中通过突变、生物化学和生物物理策略进行测试。遗传和比较生化证据表明，TrmE蛋白促进C1阻断基团的Fe/S蛋白。突变和生物化学方法将被用来测试这一想法。结构和基因组背景信息表明，APE 157蛋白通过介导不同的反应在功能上取代了关键的叶酸再循环酶5-甲酰四氢叶酸环连接酶（5-FCL）。对于原核和植物APE 157蛋白，这一假设将通过结合遗传学，代谢产物分析，同位素标记和酶学进行测试。将YgfZ表征为参与Fe/S簇代谢的普遍存在的叶酸依赖性蛋白质将是叶酸生物化学中的一个新的出发点，正如在TrmE、YgfZ和Fe/S酶之间建立叶酸连接一样。确定APE 157作为5-FCL的广泛替代品是具有普遍重要性的叶酸生物化学中未探索的方面。 更广泛的影响：尽管基因组学是一种发现原核生物和植物中未知代谢基因的强大且具有成本效益的方法，但目前尚未得到充分利用。为了促进基因组学在代谢生物化学中的应用，本项目将利用SEED数据库及其工具开发一个为期两天的研讨会。该研讨会将每年在两所主要大学举行，颁发给植物和原核生物化学家的目标群体，包括博士生，博士后和教师。除了这一更广泛的推广，该项目将有助于至少两个博士生和两个博士后在PI和Co-PI的实验室的培训，并提供本科生的研究机会。通过该项目产生的数据和信息将通过拟南芥信息资源（TAIR：www.arabidopsis.org）提供。种子库和任何独特的DNA材料将通过ABRC提供。http://www.biosci.ohio-state.edu/~plantbio/Facilities/abrc/abrchome.htm