Investigations of Protein Translocation in Archaea

古细菌中蛋白质易位的研究

• 批准号: 9816411
• 负责人: Mechthild Pohlschroder
• 金额: $ 33.3万
• 依托单位: University of Pennsylvania
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-05-01 至 2003-01-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=9816411&HistoricalAwards=false
• 关键词: Investigations; Protein; Translocation; Archaea

M. PohlschroderUniversity of Pennsylvania Protein translocation across hydrophobic membranes is an essential process in all living organisms. Biochemical and genetic analyses of the bacterial and eukaryotic systems have shown that many proteins cross these membranes via an evolutionarily conserved pore. The mechanisms and energetics seem to be distinct and much has to be learned about the roles of the components involved in this process. Sequence comparisons suggest that organisms of the third domain of life, the archaea, contain a combination of homologs of the bacterial and eukaryotic protein translocation components, suggesting that some aspects of protein translocation through the cytoplasmic membrane of these organisms are unique. However, protein translocation studies in archaea, have been limited to sequence comparisons. This research is aimed at studying an archaeal protein translocation system. The specific aims of the project are: 1. To clone and sequence archaeal homologs of known protein translocation components, and provide necessary tools to study and validate the experimental system; and 2. To use a biochemical approach and reverse genetics to purify proteins associated with the archaeal translocon. In addition to identifying H. volcanii homologs of the known bacterial and eukaryotic protein translocation machinery, this research may lead to the isolation of archaea-specific components, or components that have not been previously identified in bacteria and eukaryotes. Gaining insights into archaeal protein translocation may help us to obtain a better understanding of this conserved process all domains of life, bacteria, archaea, and eukaryotes.The ability to transport proteins across hydrophobic membranes is fundamental to the growth and survival of all cells. For example, in eukaryotes, secreted antibodies are essential for the defense against pathogens. On the other hand, many prokaryotes secrete toxins, which are central features of their pathogenicity. Thus, a better understanding of the mechanisms by which proteins traverse the membranes may allow us to modulate the secretion of pathogenic compounds as well as optimize defense mechanisms against them. Prokaryotes and lower eukaryotes also secrete enzymes to scavenge extracellular molecules. Many of these proteins have useful biotechnology applications. Understanding protein export mechanisms will help in the overexpression of recombinant secreted proteins. The archaea, which, along with bacteria and eukaryotes comprise one of the three domains of life, represent a relatively unexplored area in our study of protein translocation. An exploration of the biology of protein translocation in these organisms will not only shed light on novel processes by which proteins traverse hydrophobic archaeal membranes, but will also provide new insights into the mechanisms of previously identified bacterial and eukaryotic protein translocation systems.

M. Pohlschroder 宾夕法尼亚大学 蛋白质跨疏水膜的易位是所有生物体中的一个重要过程。对细菌和真核系统的生化和遗传分析表明，许多蛋白质通过进化上保守的孔穿过这些膜。其机制和能量学似乎是不同的，并且必须了解该过程中涉及的组件的作用。序列比较表明，生命第三域的生物体——古细菌，含有细菌和真核蛋白质易位成分的同源物组合，这表明这些生物体通过细胞质膜的蛋白质易位的某些方面是独特的。然而，古细菌中的蛋白质易位研究仅限于序列比较。本研究旨在研究古菌蛋白质易位系统。该项目的具体目标是： 1. 对已知蛋白质易位成分的古菌同源物进行克隆和测序，为研究和验证实验系统提供必要的工具； 2. 使用生化方法和反向遗传学来纯化与古菌易位子相关的蛋白质。除了鉴定已知细菌和真核生物蛋白质易位机制的火山螺杆菌同源物之外，这项研究还可能导致古细菌特异性成分或以前在细菌和真核生物中未鉴定过的成分的分离。了解古细菌蛋白质易位可能有助于我们更好地了解生命、细菌、古细菌和真核生物等所有领域的这一保守过程。跨疏水膜转运蛋白质的能力是所有细胞生长和生存的基础。例如，在真核生物中，分泌的抗体对于防御病原体至关重要。另一方面，许多原核生物分泌毒素，这是其致病性的核心特征。因此，更好地了解蛋白质穿过膜的机制可能使我们能够调节致病化合物的分泌并优化针对它们的防御机制。原核生物和低等真核生物也分泌酶来清除细胞外分子。许多这些蛋白质具有有用的生物技术应用。了解蛋白质输出机制将有助于重组分泌蛋白的过度表达。古细菌与细菌和真核生物一起构成了生命的三个领域之一，在我们的蛋白质易位研究中代表了一个相对未经探索的领域。对这些生物体中蛋白质易位生物学的探索不仅将揭示蛋白质穿过疏水性古菌膜的新过程，而且还将为先前确定的细菌和真核蛋白质易位系统的机制提供新的见解。