Amino Terminal Processing of Eukakryotic Proteins

真核蛋白的氨基末端加工

• 批准号: 9512655
• 负责人: Yie-Hwa Chang
• 金额: $ 27.5万
• 依托单位: Saint Louis University
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 1995
• 资助国家: 美国
• 起止时间: 1995-08-01 至 1999-07-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=9512655&HistoricalAwards=false
• 关键词: Amino; Terminal; Processing; Eukakryotic; Proteins

; R o o t E n t r y F 4 @ C o m p O b j b W o r d D o c u m e n t O b j e c t P o o l 4 4 F Microsoft Word 6.0 Document MSWordDoc Word.Document.6 ; Oh +' 0 $ H l D h R:\WWUSER\TEMPLATE\NORMAL.DOT marcia steinberg marcia steinberg @ qP 4 @ {\! e = e | o " " " " " " " L L L L L d n L C x | | | | | | | # ı e V T 4 " | | | | | | " " | x | | | | " | " | 6 " " " " | | 4 | 9512655 Chang PRIVATE The broad goal of this project is to understand the mechanism of amino terminal processing of eukaryotic proteins. The approach used is the purification, characterization and genetic analysis of two methionine aminopeptidases (MAPs) in Saccharomyces cerevisiae. The first aim is to elucidate the roles of yeast MAP1 in amino terminal processing and cell growth. Yeast MAP 1 contains two zinc fingers and a cobalt dependent catalytic domain. To understand the roles of MAP1 in this processing event, this investigator plans to study the roles of the zinc fingers and cobalt ions in MAP1 function. The results will show whether MAP1 is ribosome associated, and if so, how deletion of zinc fingers affects thi s interaction The P.I. also plans to compare the efficiency of methionine removal of several model proteins in yeast cells that express no MAP1, truncated MAP1 which lacks zinc fingers, and wild type MAP1 and to elucidate the roles of cobalt in MAP1 function by determining the cobalt binding constant and studying the effects of mutations at the cobalt binding sites on cell growth and enzyme activity. The second aim is to elucidate the roles of is MAP2 in N terminal processing and to determine whether it is a bifunctional protein. The P.I. plans to purify and characterize this enzyme, to compare its substrate specificity and other properties with those of MAP1, and to knock out the MAP2 gene alone and both MAPl and MAP2 genes, and characterize the phenotype of the knockout yeast strains. Amino terminal processing is one of the most common protein processing events, which occurs in all living cells and is essential for normal cell growth. Yet, until recently, very little was known about the mechanism by which this important processing event is carried out in eukaryotic cells, and how it is regulated. This study should provide novel insights into the mechanism and regulation of this important event, which in turn may lead to better understanding of the regulation of protein synthesis and the regulation of the functions of myristoylated proteins, including certain oncogene products, viral proteins, and proteins involved in transmembrane signaling and protein secretory vesicular trafficking. %%% This research project is focused on studying the mechanism of the removal of the initial amino acid from the origin of a protein called the N terminus. This is one of the most common systems for modifying newly made proteins. The generality of this phenomenon in proteins from microbes to man indicates its fundamental importance to all living organisms. Yet, relatively little is known about the mechanism by which this initial amino acid is removed, particularly in higher organisms like man, or the manner in wh ich this process is regulated. The Principal Investigator has already made considerable progress in developing the problem. He has found two enzymes which are specific for removing the N terminal methionine (the initial amino acid) from newly synthesized proteins as they are produced in eukaryotic cells. The next immediate goal of his research is to fully characterize these interesting proteins, so called methionine aminopeptidases (MAPs), that are important to every living cell. It is certain that understanding the novel features of yeast MAPs will provide new insights into the mechanism and the regulation of this cellular event that is essential for cell growth. *** @ ....()()))()() Z : phoenix W EH o ; S u m m a r y I n f o r m a t i o n ( 4 @ E 4 @ Microsoft Word 6.0 4

;​R o o o o o o o o o o o o o o o o o o o o o o o o o o o o o o o o o o o o o o o o o o o o o o o o o o o o o​F Microsoft Word 6.0文档MSWordDoc。6;Oh +' 0 $ H l D H R:\WWUSER\TEMPLATE\NORMAL。DOT玛西亚·斯坦伯格玛西亚·斯坦伯格@ qp4 @ {\！e = e | 0 " " " " " " " L L L L L L L d n L C x | | | | | | | # ye V T 4 " | | | | | | " " |x | | | | " | " | 6 " " " " | | 4 | 9512655 Chang PRIVATE本项目的主要目标是了解真核蛋白氨基末端加工的机制。本文采用的方法是对酿酒酵母中两种蛋氨酸氨基肽酶（MAPs）进行纯化、鉴定和遗传分析。第一个目的是阐明酵母MAP1在氨基末端加工和细胞生长中的作用。酵母map1含有两个锌指和一个依赖钴的催化结构域。为了了解MAP1在这一加工过程中的作用，本研究者计划研究锌指和钴离子在MAP1功能中的作用。结果将显示MAP1是否与核糖体相关，如果是的话，锌指的缺失如何影响这种相互作用。P.I.还计划比较几种模型蛋白在酵母细胞中去除蛋氨酸的效率，这些模型蛋白不表达MAP1，缺少锌指的截断MAP1，并通过测定钴结合常数和研究钴结合位点突变对细胞生长和酶活性的影响来阐明钴在MAP1功能中的作用。第二个目的是阐明MAP2在N末端加工中的作用，并确定它是否是一个双功能蛋白。P.I.计划对该酶进行纯化和表征，将其底物特异性和其他特性与MAP1进行比较，并单独敲除MAP2基因以及敲除MAPl和MAP2基因，并对敲除酵母菌株的表型进行表征。氨基末端加工是最常见的蛋白质加工事件之一，发生在所有活细胞中，是正常细胞生长所必需的。然而，直到最近，人们对真核细胞中这一重要加工过程的机制以及它是如何被调节的知之甚少。这项研究将为这一重要事件的机制和调控提供新的见解，进而可能导致更好地理解蛋白质合成的调控和肉豆蔻酰基化蛋白的功能调控，包括某些致癌基因产物、病毒蛋白、参与跨膜信号传导和蛋白分泌囊泡运输的蛋白。这个研究项目的重点是研究从一种叫做N端的蛋白质的起源中去除初始氨基酸的机制。这是一种最常见的修饰新合成蛋白质的系统。这种现象在从微生物到人类的蛋白质中的普遍性表明了它对所有生物体的根本重要性。然而，对于这种初始氨基酸被移除的机制，特别是在像人类这样的高等生物中，或者这个过程是如何被调节的，人们所知相对较少。首席研究员在解决这个问题方面已经取得了相当大的进展。他发现了两种酶，当真核细胞中新合成的蛋白质产生时，它们可以特异性地去除N端蛋氨酸（初始氨基酸）。他研究的下一个直接目标是充分表征这些有趣的蛋白质，即所谓的蛋氨酸氨基肽酶（MAPs），它们对每个活细胞都很重要。可以肯定的是，了解酵母MAPs的新特征将为细胞生长所必需的细胞事件的机制和调控提供新的见解。*** @ ....（）（）) ()() ()() Z：凤凰W ho；如果我是你，我是你，我是你，我是你，我是你。Microsoft Word 6.0