Tracing the Origins of the Chaperonin (CCT) Complex in Eukaryotes

通过最近的宏基因组分析，真核生物的根源已经变得很清楚，在古细菌王国的微生物。该项目将探索一个古老的蛋白质家族的起源，帮助蛋白质折叠和稳定性，伴侣蛋白（cpn），跟踪他们的进化到真核生物。最近在所谓的阿斯加德古菌基因组中发现的CPN编码基因将被利用，以获得这些蛋白质的生化功能的结构见解，这些蛋白质以前由于生物体的不可培养性而无法获得。该项目将利用来自宏基因组组装基因组（MAGs）的基因合成和蛋白质表达。主要的科学效益将是通过应用合成生物学来深入了解它们的祖先蛋白质折叠功能和真核生物从原核生物祖先的发展。适当的蛋白质折叠对所有生命过程都至关重要，对健康和疾病都有影响。更广泛的影响将通过培训一名工作人员、科学家和本科生，以及通过外联活动。该项目的主要目标是在新发现的CPN60系统发育树最深分支中发现蛋白质功能的定量生化见解。快速积累的基因组序列和来自古细菌最深处分支的单扩增基因组（SAGS）揭示了基因组复杂性和基因复制增加的谱系。研究人员已经在ASGARD古细菌中发现了不同的CPN60基因，这些基因似乎是现存真核生物中发现的蛋白质的CCT亚基的祖先。来自古细菌和真核生物的CPN60对专性折叠底物（如微管蛋白）均表现出合作变构效应。为了探索真核CCT复合物的功能进展，本工作将从ATP结合、ATP酶活性和蛋白质折叠能力方面评估这些古老的CPN基因和复合物伴侣的功能。因为在阿斯加德古菌单扩增基因组中也有微管蛋白同源物，该项目将解决一个令人兴奋的假设，即古菌蛋白酶静止是真核生物中发现的详细的伴侣/客户蛋白相互作用网络的祖先试验场。并将追踪伴侣复合物从具有基本生化特征的假定祖先到现代真核生物共有的高度特化和分化的真核CCT复合物的功能进展。该奖项反映了美国国家科学基金会的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准进行评估，被认为值得支持。

Through recent metagenomic analysis, it has become clear that the root of eukaryotes lies among microbes in the kingdom archaea. This project will explore the origins of an ancient protein family that assists with protein folding and stability, the chaperones (CPNs), to track their evolution into the eukaryotes. Recent identification of CPN encoding genes within the genomes of the so-called Asgard archaea will be utilized in order to gain structural insights into the biochemical function of these proteins, which have previously not been available due to the unculturability of the organisms. The project will utilize gene synthesis and expression of protein from metagenomic assembled genomes (MAGs). The main scientific benefit will be through the application of synthetic biology to gain insights into their ancestral protein folding functions and the development of eukaryotes from their prokaryotic ancestors. Proper protein folding is essential to all life processes and has implications in health and disease. Broader impacts will be through the training of a staff scientist and and undergraduates, as well as through outreach activities. The main goal of the project is the discovery of quantitative biochemical insights into the functions of proteins in the newly discovered deepest branch of the CPN60 phylogenetic tree. Rapidly accumulating genome sequences and Single Amplified Genomes (SAGS) from the deepest branches of the archaea reveal lineages with increases in genome complexity and gene duplication. The investigators have identified divergent CPN60 genes in the ASGARD archaea which appear to be ancestral to the CCT subunits of the proteins found in extant eukarya. CPN60 from both the archaea and eukaryotes show cooperative allosteric effects in response to an obligate folding substrate such as tubulin. To explore the functional progression towards the eukaryotic CCT complex, this work will assess the function of these ancient CPN genes and the complex chaperones in terms of their ATP binding and ATPase activity and protein folding ability. Because there are also tubulin homologs in the Asgard Archaea Single Amplified Genomes, the project will address the exciting hypothesis that archaeal proteostasis was the ancestral proving ground for the detailed chaperone/client protein interaction networks found in eukaryotes, and will trace the functional advances of the chaperone complex from a putative ancestor with rudimentary biochemical features to the highly specialized and differentiated eukaryotic CCT complex shared by modern eukaryotes.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.