A Functional Genomics Study of the Primary Pathways for Carbon Metabolism in a Hyperthermophilic Archaeon

超嗜热古菌碳代谢主要途径的功能基因组学研究

• 批准号: 0129841
• 负责人: Michael Adams
• 金额: $ 48万
• 依托单位: University of Georgia Research Foundation Inc
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-06-01 至 2006-05-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=0129841&HistoricalAwards=false
• 关键词: Functional; Genomics; Study; Primary; Pathways

Well over 200 genomic sequencing efforts have been completed ranging from pathogens and exotic microbes, to yeast, rice and human, yielding a phenomenal array of sequence information, the comprehension of which is only just beginning. Understanding how a living cell works, however, obviously requires much more than simply determining the genome sequence and the number of genes. A fundamental problem is how to find genes and assign functions to these genes. Insights can be obtained by analyzing the sequences of the proteins that the genes encode using the available databases, but by such methods less than half of the genes in a given organism can be assigned a function with any degree of confidence. Hence, the roles of about half of the genes in any given genome is essentially unknown. This suggests that half of life's biochemistry is unknown. A major effort is therefore needed to assign biological roles to these unknown genes through functional genomic approaches.The goal of this research is to identify genes involved in carbon metabolism of Pyrococcus furiosus, an archaeon (archaebacterium) that grows optimally at 100C. The research will largely focus on finding genes involved in carbon metabolism in the 'unknown' half of the microbial genome. The P. furiosus 1.908 Mb genome contains approximately 2,200 putative genes. The fundamental hypothesis to be tested is that many of the more than 1,000 genes of unknown function play key roles in primary carbon metabolism by as yet unknown mechanisms. This will be ascertained using DNA microarray analyses using all 2,200 genes in the P. furiosus genome. Preliminary data have shown that a surprisingly large number of the unknown genes are regulated by the carbon source used to grow the organism. The specific aims are a) to identify previously uncharacterized genes involved in the metabolism of carbohydrates, proteins and C-1 compounds by analyzing expression levels using cells grown under a wide variety of conditions, b) to confirm gene expression data by enzymatic assays and metabolite analyses, and c) to characterize the key enzymes by direct purification using P. furiosus biomass from 600 liter fermentations and by using recombinant proteins that are available through a complementary structural genomics effort with P. furiosus. It is anticipated that the results of this study will provide completely new insights into the metabolism of fixed carbon compounds and the role of what are currently hypothetical genes in both P. furiosus and in other organisms, including human.

已经完成了200多个基因组测序工作，从病原体和外来微生物到酵母、水稻和人类，产生了大量序列信息，对这些信息的理解才刚刚开始。 然而，要了解活细胞是如何工作的，显然需要的不仅仅是确定基因组序列和基因数量。 一个基本的问题是如何找到基因和分配功能给这些基因。 通过分析基因编码的蛋白质序列，可以利用现有的数据库获得洞察力，但是通过这种方法，在给定的生物体中，只有不到一半的基因可以被赋予任何程度的置信度。 因此，在任何给定的基因组中，大约一半的基因的作用基本上是未知的。 这表明生命的一半生物化学是未知的。 因此，需要通过功能基因组学方法来分配这些未知基因的生物学作用。本研究的目标是确定参与激烈火球菌碳代谢的基因，激烈火球菌是一种在100 ℃下生长最佳的古细菌。 这项研究将主要集中在寻找微生物基因组中“未知”的那一半中参与碳代谢的基因。 P. furiosus 1.908 Mb基因组包含大约2，200个推定基因。 有待检验的基本假设是，在1,000多个功能未知的基因中，有许多基因通过迄今未知的机制在初级碳代谢中发挥关键作用。 这将通过使用P. furiosus基因组中所有2，200个基因的DNA微阵列分析来确定。 初步数据表明，令人惊讶的是，大量的未知基因受到用于生长生物体的碳源的调控。 具体目的是a）通过使用在多种条件下生长的细胞分析表达水平来鉴定参与碳水化合物、蛋白质和C-1化合物代谢的先前未表征的基因，B）通过酶测定和代谢物分析来确认基因表达数据，以及c）通过使用来自600升发酵的P. furiosus生物质直接纯化和通过使用可通过以下途径获得的重组蛋白来表征关键酶：互补结构基因组学研究。 预计这项研究的结果将为固定碳化合物的代谢以及目前假设的基因在P. furiosus和其他生物（包括人类）中的作用提供全新的见解。