Pyridoxal 5'-phosphate homeostasis in Escherichia coli

大肠杆菌中吡哆醛 5-磷酸稳态

• 批准号: 10439656
• 负责人: Valerie A de Crecy-Lagard
• 金额: $ 29.84万
• 依托单位: UNIVERSITY OF FLORIDA
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-09-01 至 2024-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10439656
• 关键词: Address; Affect; Animal Model; Anti-Bacterial Agents; Antiparasitic Agents; Area; Bacillus subtilis; Bacteria; Binding; Biochemical; Chemicals; Cyanobacterium; Databases; Defect; Disease; Enzymes; Escherichia coli; Escherichia coli K12; Eukaryotic Cell; Family; Family member; Genes; Genetic; Genomic approach; Genomics; Goals; Homeostasis; Human; Laboratories; Life; Link; Mammals; Manuscripts; Metabolic; Metabolic Pathway; Metabolism; Methods; Mission; Mitochondria; Molecular; Nervous system structure; Operon; Organism; Orphan; Orthologous Gene; Pathogenesis; Pathology; Pathway interactions; Phosphate-Binding Proteins; Plants; Process; Protein Family; Proteins; Publishing; Pyridoxal Phosphate; Reaction; Regulation; Research; Research Support; Role; Site; Study models; Systems Biology; Testing; United States National Institutes of Health; Vitamin B6; Vitamins; Work; cofactor; comparative genomics; differential expression; human disease; inhibitor; member; mutant; nervous system disorder; pathogenic bacteria; response; transcriptome sequencing; transcriptomics

One of the major and underappreciated issues of the post-genomic era is that the precise molecular functions of half of the proteins in databases are not known. Even in Escherichia coli K12 MG1655 one of the most studied model organism, ~1200 genes are still of unknown (or erroneous) function. In parallel, open questions remain in metabolic areas that were thought to be well studied like vitamin synthesis and salvage. For example, vitamin B6 in its active form as pyridoxal 5'-phosphate (PLP) is a cofactor for over 180 PLP-dependent enzymes that are involved in many metabolic pathways. PLP is a highly reactive molecule and is both labile and toxic so the intracellular concentration of PLP not bound to proteins remains low. How PLP is delivered to target enzymes in these conditions remains a mystery in all organisms. In addition, while the core E. coli B6 synthesis pathway is well described, the transporter(s) involved in salvage are unidentified and very little is known about regulation of PLP synthesis/salvage genes. This project focuses on identifying “missing” players in PLP metabolism in E. coli. In this process, the functions of several orphan E. coli genes should be solved and, more generally, key conserved players in PLP homeostasis characterized. Preliminary results published in two manuscripts have shown that the E. coli YggS protein, a member of the COG0325 as well as its human ortholog PROSC that has been linked to PLP-dependent diseases are involved in PLP homeostasis. The goal of Aim 1 is to test the hypothesis that this family could be the “missing” key player in delivering the PLP cofactor to target enzymes. Aim 2 focuses the response to PLP imbalance by identifying missing E. coli B6 transporters and characterizing the response to high Pyridoxin levels. The final exploratory aim will focus on two other orphan PLP binding proteins and an orphan PLP-binding regulator. The proposed research is significant because in humans, toxic levels of PLP or deficiency of PLP or defects affecting PLP metabolism are linked to many pathologies, particularly of the nervous system, hence understanding how PLP is delivered to target enzymes will have implications for a wide range of human diseases. Also, many enzymes of PLP metabolism have been recognized as antibacterial and antiparasitic targets, thus understanding PLP homeostasis is critical to develop successful inhibitors. Finally, this work should characterize 5 genes of unknown functions in E. coli, a major model organism.

后基因组时代的主要且未被充分认识的问题之一是，精确的 数据库中一半蛋白质的分子功能尚不清楚。即使在大肠杆菌中 K12 MG1655 是研究最多的模式生物之一，约 1200 个基因仍然未知（或 错误）函数。与此同时，代谢领域仍然存在悬而未决的问题，这些问题被认为是 像维生素合成和回收一样得到充分研究。例如，活性形式的维生素 B6 吡哆醛 5'-磷酸 (PLP) 是 180 多种 PLP 依赖性酶的辅助因子 在许多代谢途径中。 PLP 是一种高反应性分子，不稳定且有毒，因此 未与蛋白质结合的 PLP 的细胞内浓度仍然很低。 PLP 如何交付给 在所有生物体中，这些条件下的目标酶仍然是一个谜。此外，虽然核心 大肠杆菌 B6 合成途径已得到充分描述，参与回收的转运蛋白是 PLP 合成/挽救基因的调控尚未被识别且知之甚少。这个项目 专注于识别大肠杆菌 PLP 代谢中“缺失”的参与者。在此过程中，函数 应该解决几个孤儿大肠杆菌基因的问题，更一般地说，解决关键保守基因的问题 PLP 稳态特征。两篇手稿发表的初步结果表明 大肠杆菌 YggS 蛋白，COG0325 的成员及其人类直系同源物 PROSC 与 PLP 依赖性疾病相关的 PLP 稳态也参与其中。目标是 目标 1 是检验这一假设：这个家庭可能是实现这一目标的“缺失”关键角色。 PLP 辅助因子靶向酶。目标 2 通过识别来重点应对 PLP 失衡 缺失大肠杆菌 B6 转运蛋白并表征对高吡哆醇水平的反应。决赛 探索性目标将集中于另外两个孤儿 PLP 结合蛋白和一个孤儿 PLP 结合蛋白 调节器。拟议的研究意义重大，因为在人类中，PLP 或 PLP 缺乏或影响 PLP 代谢的缺陷与许多病理学相关，特别是 神经系统，因此了解 PLP 如何传递到目标酶将有 对多种人类疾病的影响。此外，PLP 代谢的许多酶具有 被认为是抗菌和抗寄生虫靶标，从而了解 PLP 稳态 对于开发成功的抑制剂至关重要。最后，这项工作应该表征 5 个基因 大肠杆菌（一种主要的模式生物）中的未知功能。

项目成果

Pyridoxal 5'-phosphate synthesis and salvage in Bacteria and Archaea: predicting pathway variant distributions and holes.

• DOI: 10.1099/mgen.0.000926
• 发表时间: 2023-03
• 期刊: MICROBIAL GENOMICS
• 影响因子: 3.9
• 作者: Denise, Remi;Babor, Jill;Gerlt, John A.;de Crecy-Lagard, Valerie
• 通讯作者: de Crecy-Lagard, Valerie

Uncovering the chemistry of C-C bond formation in C-nucleoside biosynthesis: crystal structure of a C-glycoside synthase/PRPP complex.

• DOI: 10.1039/d0cc02834g
• 发表时间: 2020-07-14
• 期刊: Chemical communications (Cambridge, England)
• 作者: Gao S;Radadiya A;Li W;Liu H;Zhu W;de Crécy-Lagard V;Richards NGJ;Naismith JH
• 通讯作者: Naismith JH