Functional Genomics of D-Amino Acid Metabolism in Pseudomonas Aeruginosa

铜绿假单胞菌 D-氨基酸代谢的功能基因组学

• 批准号: 0950217
• 负责人: Chung-Dar Lu
• 金额: $ 64万
• 依托单位: Georgia State University Research Foundation, Inc.
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-03-01 至 2015-12-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=0950217&HistoricalAwards=false
• 关键词: Functional; Genomics; Amino; Acid; Metabolism

Intellectual MeritD-amino acids are naturally synthesized in all living organisms to serve in a variety of specific functions. Examples include the well-known D-Ala and D-Glu as essential components of bacterial cell wall to the latest discovery of D-Ser as neurotransmitter in humans. The level of D-amino acids inside the cells is subjected to tight regulation to prevent any potential harmful effects. Up to date, the biochemistry of D-amino acid catabolism has not been intensively studied in comparison to those of L-amino acids, and therefore, the information is still fragmentary. Famous for its enormous catabolic capacity, P. aeruginosa is able to utilize many D-amino acids as nutrient, and hence serves as an excellent model organism to explore novel pathways and enzymes for D-amino acid metabolism. It is best evidenced by a recent report of a new type of D-to-L arginine racemization by coupled catabolic and anabolic dehydrogenases discovered by the PI's group on a previous NSF-funded project. The results of preliminary studies also uncovered several important features in D-amino acid utilization. D-amino acids have two potential physiological functions - inversion to L-amino acids for peptide synthesis or degradation to serve as nitrogen or carbon source. How each of these D-amino acids can be utilized depends on the presence as well as the induction level of specific racemases or catabolic enzymes/pathways. The goal of this research is to continue efforts in exploration of metabolic pathways for D-amino acids with a systemic approach. The specific aims of this project are: (i) to elucidate physiological functions and biochemical properties of DadAX and DauAB in D-amino acid deamination and racemization; and (ii) to explore catabolic pathways for D-glutamate/glutamine/asparagine and D-proline/hydroxyproline. Experiments will be conducted to test the proposed physiological and biochemical functions of thus identified genes and their encoding enzymes. Meanwhile, a more systemic transcriptome analysis will be conducted to have a better understanding of cellular responses to the presence of these D-amino acids. Genes and encoding enzymes thus identified from this study will be subjected to further characterization accordingly regarding physiological functions, genetic regulation, and biochemical properties in D-amino acid catabolism. Up till now, only limited effort has been devoted systemically to explore the biochemical diversity toward D-amino acids. Findings from this research are anticipated to further extend the current knowledge of amino acid metabolism.Broader ImpactsThe PI plans to continue the effort in maintaining an active research environment composed of post-doc associates, MS and PhD graduate students, and undergraduate students. In addition, the PI will extend the effort for recruiting promising minority students to research by serving as faculty mentor for the McNair Program in the summers. Scientifically, the PI lab will provide unique training opportunities with a combination of modern genomics/proteomics and conventional genetics/biochemistry approaches in the area of bacterial physiology and biochemistry. In addition, the ongoing collaborations between PI and colleagues in protein crystallography and mechanistic enzymology will be fostered to benefit trainees of different disciplines, by way of regular joint group meetings and encouragement of students taking courses offered by collaborating colleagues. The success of this proposed research will have broader impacts in training competitive researchers with multi-disciplinary coverage and in recruiting under-representative minority students into careers of biological research. All members of the Lu laboratory will participate in local, national, and international research symposia to disseminate the research and educational activities of the laboratory to a broader scientific community.

智力优势D-氨基酸在所有活着的有机体中都是天然合成的，具有各种特定的功能。例子包括众所周知的D-丙氨酸和D-谷氨酸作为细菌细胞壁的基本成分，以及最新在人类中发现的D-丝氨酸作为神经递质。细胞内的D-氨基酸水平受到严格的调控，以防止任何潜在的有害影响。迄今为止，与L氨基酸相比，D-氨基酸分解代谢的生物化学还没有得到深入的研究，因此，关于D-氨基酸分解代谢的信息还很零碎。铜绿假单胞菌以其巨大的分解代谢能力而闻名，能够利用多种D-氨基酸作为营养物质，因此是探索D-氨基酸代谢新途径和酶的优秀模式生物。最近的一份报告就是最好的证据，该报告表明，在之前的一个国家科学基金会资助的项目中，PI的小组发现了一种新型的D-L精氨酸外消旋作用，它是由分解代谢和合成代谢脱氢酶偶联而成的。初步研究结果还揭示了D-氨基酸利用的几个重要特征。D-氨基酸具有两种潜在的生理功能--转化为L--作为氮源或碳源的氨基酸合成或降解。如何利用这些D-氨基酸取决于特定外消旋酶或分解代谢酶/途径的存在和诱导水平。本研究的目标是用系统的方法继续努力探索D-氨基酸的代谢途径。本项目的具体目标是：(1)阐明DadAX和DauAB在D-氨基酸脱氨和消旋过程中的生理功能和生化特性；(2)探索D-谷氨酸/谷氨酰胺/天冬酰胺和D-脯氨酸/羟脯氨酸的分解代谢途径。将进行实验，以测试所确定的基因及其编码酶的拟议生理和生化功能。同时，将进行更系统的转录组分析，以更好地了解细胞对这些D-氨基酸的反应。本研究确定的基因和编码酶将在D-氨基酸分解代谢的生理功能、遗传调控和生化特性方面进行进一步的表征。到目前为止，系统地探索D-氨基酸的生物化学多样性的工作还很有限。这项研究的结果有望进一步扩展目前对氨基酸代谢的知识。广泛的影响PI计划继续努力维持一个由博士后助理、硕士和博士研究生以及本科生组成的活跃的研究环境。此外，PI将通过在夏季担任McNair项目的教师导师来扩大招募有前途的少数族裔学生进行研究的努力。科学上，PI实验室将提供独特的培训机会，将现代基因组学/蛋白质组学与细菌生理和生化领域的传统遗传学/生物化学方法相结合。此外，通过定期联合小组会议和鼓励学生参加合作同事提供的课程，将促进PI和同事在蛋白质结晶学和机械酶学方面的持续合作，使不同学科的学员受益。这项拟议研究的成功将在培养具有多学科覆盖面的有竞争力的研究人员和招募不具代表性的少数族裔学生从事生物研究方面产生更广泛的影响。鲁实验室的所有成员将参加地方、国家和国际研究研讨会，向更广泛的科学界传播实验室的研究和教育活动。