DOMAIN ANALYSIS OF M ACEIVORANS REPLICATION PROTEIN A 1

M ACEIVORANS 复制蛋白 A 1 的结构域分析

• 批准号: 7357984
• 负责人: ISAAC CANN
• 金额: $ 0.24万
• 依托单位: UNIVERSITY OF CALIFORNIA-IRVINE
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-08-29 至 2007-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7357984
• 关键词: Archaea; DNA; DNA binding protein; protein structure; proteins

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. The oligonucleotide/oligosaccharide-binding (OB) fold, a module ranging from 70-150 amino acids, is central to the architecture of single-stranded DNA-binding proteins. Single-stranded DNA-binding proteins are essential in diverse cellular processes. The bacterial single-stranded DNA-binding protein, a single polypeptide known as SSB, harbors a single OB fold. In contrast, the eukaryotic functional homolog, known as replication protein A (RPA), is a heterotrimeric protein containing multiple OB folds. In the methanogenic archaea, single polypeptide RPA proteins with multiple OB folds and a zinc finger domain have been described. The OB folds of these proteins were more similar to their eukaryotic counterparts than the bacterial ones. Here, we describe the functional analysis of a new form of RPA from the methanogenic archaeon Methanosarcina acetivorans. The novel RPA, designated MacRPA1, is comprised of four OB folds and lacks the zinc finger domain hitherto found in the RPA proteins of methanogens. MacRPA1 was compared with two other RPA proteins in M. acetivorans for their effects on DNA strand exchange, DNA synthesis, and cleavage of a flap by cognate proteins involved in these processes. MacRPA1 stimulated DNA strand exchange and primer extension reactions, however, it suppressed cleavage of a flap by the methanosarcinal homolog of flap endonuclease 1. N-terminal and C-terminal truncated derivatives of MacRPA1 were made and their properties were studied using biochemical and biophysical methods. Most interestingly, comparison of MacRPA1 with potential orthologs provided critical insight into a process that may serve to generate new OB folds in cells.

该子项目是利用NIH/NCRR资助的中心赠款提供的资源的许多研究子项目之一。子弹和调查员（PI）可能已经从其他NIH来源获得了主要资金，因此可以在其他清晰的条目中代表。列出的机构适用于该中心，这不一定是调查员的机构。寡核苷酸/寡糖结合（OB）折叠，范围从70-150氨基酸的模块是单链DNA结合蛋白的构建的核心。单链DNA结合蛋白在各种细胞过程中至关重要。细菌的单链DNA结合蛋白（一种称为SSB）具有单个OB折叠。相比之下，真核功能同源物（称为复制蛋白A（RPA））是包含多个OB折叠的异三聚体蛋白。在甲烷古细菌中，已经描述了具有多个OB折叠和锌指域的单多肽RPA蛋白。这些蛋白质的OB褶皱比细菌的蛋白质与真核对应物更相似。在这里，我们描述了来自甲烷元素甲氧甲菌乙烷的新形式RPA的功能分析。新颖的RPA被指定为MACRPA1，由四个OB折叠组成，缺少迄今为止在甲烷元素RPA蛋白中发现的锌指域。将MACRPA1与M. acetivorans中的另外两种RPA蛋白进行了比较，从而对DNA链交换，DNA合成以及涉及这些过程中的同源蛋白的皮瓣裂解的影响。然而，麦克比1（MACRPA1）刺激的DNA链交换和引物延伸反应，它抑制了通过皮瓣核酸内核酶的甲壳同源物1。N-末端和C末端截短的麦克帕尔衍生物的裂纹，并使用生物化学和生物物理学进行了研究。最有趣的是，MACRPA1与潜在的直系同源物的比较为可能产生细胞中新的OB折叠的过程提供了重要的见解。