SGER: A Novel Protein-linked Route for the Biosynthesis of Indole-3-acetic Acid in Maize Endosperm

SGER：玉米胚乳中吲哚-3-乙酸生物合成的新型蛋白连接途径

• 批准号: 0643845
• 负责人: Jerry Cohen
• 金额: --
• 依托单位: University of Minnesota-Twin Cities
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-09-01 至 2008-02-29
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=0643845&HistoricalAwards=false
• 关键词: SGER; Novel; Protein; linked; Route

The biosynthesis of the primary auxin-type plant hormone, indole-3-acetic acid (IAA), has remained an active area of study for more than half a century and numerous different pathways have been predicted but few have been shown to be critical for normal plant development. The biosynthesis of IAA during late kernel development in maize is important for the accumulation of relatively large quantities of stored conjugated IAA and has been shown to originate from the amino acid tryptophan. The physiologically relevant production of high levels of IAA during a short time period in endosperm development makes this a unique system for analysis of IAA biosynthesis pathways. Using two complementary approaches to identify the enzymes responsible for the different conversions in the pathway as well as the identification of the intermediate compounds, it was shown that previously reported pathways could not account for the measured IAA production. A metabolite screen for potential intermediates revealed the majority of the radioactivity that was not either tryptophan or IAA was associated with a higher molecular weight fraction. Gel electrophoresis and size exclusion HPLC indicated radioactivity was associated with proteins in the size range of 10-20 kDa, where tryptophan was linked apparently via a thio-ester bond. The focus of this project is to utilize biochemical analyses to isolate, characterize and microsequence proteins, which become bound to tryptophan, for subsequent cloning of corresponding genes. The ability to produce the tryptophyl-proteins in larger amounts and at higher purities will help resolve their role(s) in IAA biosynthesis.Broader impacts: This project will train a postdoctoral scientist in both research and in the grant writing process. Results obtained in this project will be used as part of a separately funded effort to train and mentor minority undergraduate students. In addition, the laboratory will host visiting students, postdocs and other scientists from universities and 4-year colleges to learn techniques related to the project goals.

生长素类植物激素吲哚-3-乙酸（IAA）的生物合成在半个多世纪以来一直是一个活跃的研究领域，并且已经预测了许多不同的途径，但很少被证明对正常植物发育至关重要。玉米籽粒发育后期IAA的生物合成对于积累相对大量的储存共轭IAA是重要的，并且已显示其来源于氨基酸色氨酸。在胚乳发育的短时间内生理相关地产生高水平的IAA，使其成为分析IAA生物合成途径的独特系统。使用两种互补的方法来确定负责不同的转换途径中的酶，以及中间化合物的鉴定，它表明，以前报道的途径不能占测得的IAA生产。 潜在中间体的代谢物筛选显示，大部分非色氨酸或IAA的放射性与较高分子量组分相关。凝胶电泳和分子排阻HPLC表明放射性与10-20 kDa范围内的蛋白质相关，其中色氨酸显然通过硫酯键连接。 该项目的重点是利用生物化学分析来分离、表征和微测序与色氨酸结合的蛋白质，以便随后克隆相应的基因。 以更大的数量和更高的纯度生产蛋白质的能力将有助于解决它们在IAA生物合成中的作用。更广泛的影响：该项目将在研究和资助写作过程中培养博士后科学家。在这个项目中获得的成果将被用作一个单独资助的努力，以培训和指导少数民族本科生的一部分。 此外，实验室将接待来自大学和四年制学院的访问学生、博士后和其他科学家，学习与项目目标相关的技术。