MRI: Acquisition of a Phosphorimager to Investigate Phosphorylation and Regulation of TACE

MRI：获取磷成像仪来研究 TACE 的磷酸化和调节

• 批准号: 0215896
• 负责人: Blaine Moore
• 金额: $ 7.39万
• 依托单位: Kalamazoo College
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-08-01 至 2005-07-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=0215896&HistoricalAwards=false
• 关键词: MRI; Acquisition; Phosphorimager; Investigate; Phosphorylation

A grant has been awarded to Kalamazoo College under the supervision of Dr. Moore,to acquire a phosphoimager, which will allow faculty to investigate phosphorylation and regulation of the TNF-alpha converting enzyme (TACE). TACE is a protein that works as an enzyme, catalyzing the cleavage of large, full-length precursors (known as substrates of TACE) into smaller fragments. Understanding how the activity of the TACE enzyme is regulated will provide insights as to how, mechanistically, these fragments are generated. Specific experiments are planned to determine the sites (specific amino acids) on TACE that are phosphorylated by PKC, and to determine whether the phosphorylation state of TACE directly influences its activity. The former involves the use of mutated versions of TACE that lack critical proposed phosphorylation sites. This experiment will determine where on TACE these phosphate groups are added. The latter also involves the use of the aforementioned mutant versions of TACE in cleavage assays to determine whether the cleavage of TACE substrates (other proteins that TACE acts on and cleaves) is affected by phosphorylation state. In addition to providing basic science information about the regulation of the TACE enzyme, the experiments described above will have a much broader impact. For example, much of what we learn about the TACE enzyme may be generalized to similar proteases that are in the same protein "family" as TACE. Furthermore, understanding the mechanisms whereby TACE is "turned on" will illuminate general strategies used by cells to "turn on" protein enzymes. The broader impact of this work lies in uncovering the complexities of cellular function: the more scientist know about how cells work normally, the better prepared scientists are to determine what goes wrong in malfunctioning cells in disease or injury states. Moreover, because these experiments will be done in conjunction with undergraduate student collaborators, an important educational experience will be provided. In addition several other faculty members in the college are anxious to learn application of the technique, and they will benefit from the presence of this piece of equipment on the campus. Both faculty and students from the biology and chemistry departments will be involved in research utilizing the equipment.

在摩尔博士的监督下，卡拉马祖学院获得了一笔赠款，用于购买一台磷光仪，这将使教职员工能够研究肿瘤坏死因子-α转换酶(TACE)的磷酸化和调节。TACE是一种蛋白质，作为一种酶发挥作用，催化大的全长前体(称为TACE的底物)裂解成较小的片段。了解TACE酶的活性是如何调节的，将有助于深入了解这些片段是如何从机制上产生的。计划进行具体的实验，以确定被PKC磷酸化的TACE上的位点(特定氨基酸)，并确定TACE的磷酸化状态是否直接影响其活性。前者涉及使用缺乏关键建议的磷酸化位点的突变版本的TACE。这项实验将确定这些磷酸基团在TACE上的添加位置。后者还涉及在切割分析中使用上述TACE的突变版本，以确定TACE底物(TACE作用和切割的其他蛋白质)的切割是否受磷酸化状态的影响。除了提供有关TACE酶调节的基本科学信息外，上述实验还将产生更广泛的影响。例如，我们所了解的许多关于TACE酶的知识可以推广到与TACE属于同一蛋白质“家族”的类似的蛋白酶。此外，了解TACE被“启动”的机制将有助于阐明细胞用于“启动”蛋白质酶的一般策略。这项工作的更广泛影响在于揭示了细胞功能的复杂性：科学家对细胞如何正常工作了解得越多，科学家就越有准备，以确定疾病或损伤状态下出现故障的细胞出了什么问题。此外，由于这些实验将与本科生合作者一起完成，因此将提供重要的教育经验。此外，学院中的其他几名教职员工也渴望学习这项技术的应用，他们将受益于校园内这台设备的存在。来自生物和化学系的教职员工和学生都将参与利用这些设备进行的研究。