Anion Exchange in Bacteria

细菌中的阴离子交换

• 批准号: 8905130
• 负责人: Peter Maloney
• 金额: $ 19.5万
• 依托单位: Johns Hopkins University
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 1989
• 资助国家: 美国
• 起止时间: 1989-09-01 至 1993-08-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=8905130&HistoricalAwards=false
• 关键词: Anion; Exchange; Bacteria

This research examines the physical and cellular biochemistry of a novel exchange protein found in the gram-negative anaerobe Oxalobacter formigenes. In this anaerobe, the exchange of divalent oxalate for monovalent formate forms the basis for generation of proton-motive force. This antiport reaction is unprecedented among "chemiosmotic" carriers for its role in cellular biochemistry and unique among anion exchange proteins for its electrogenic character. Assays have been devised to reconstitute this antiport reaction without loss of function and are used as a guide to purify the exchange protein from membranes of O. formigenes. After the protein is purified experiments will reveal the kinetic reaction mechanism (Ping Pong or Simultaneous) and that portion of the reaction cycle which carries charge over the membrane. Using residue-specific chemical probes, monoclonal antibodies directed against the antiport protein, and the derived amino acid sequence predictions will be made about the physical structure. Oxalobacter formigenes, a gram-negative anaerobe, is a cell whose sole source of metabolic energy derives from the decarboxylation of oxalic acid. It has been established that membranes of O. formigenes contain a carrier that displays the electrogenic exchange of divalent oxalate and monovalent formate. This reaction is of particular interest to cellular biochemistry as a mechanism whereby the organism sustains a proton-motive force. Thus, the antiport of precursor and product results in the net entry of negative charge, while consumption of a proton during oxalate decarboxylation changes internal pH. This is a novel use for such a "secondary" carrier and is of interest to determine whether other cells that utilize carboxylic acids also exploit such antiport reactions. This research will examine the molecular properties of this carrier and its overall function in cellular biochemistry. This work will contribute to the current understanding of how bacteria organize their cellular functions.

这项研究检查了革兰氏阴性厌氧菌产酸草酸杆菌中发现的新型交换蛋白的物理和细胞生物化学。 在这种厌氧菌中，二价草酸盐与一价甲酸盐的交换形成了产生质子动力的基础。 这种反向转运反应因其在细胞生物化学中的作用而在“化学渗透”载体中是前所未有的，并且因其产电特性而在阴离子交换蛋白中是独一无二的。 已设计了一些测定来重建这种反向转运反应而不丧失功能，并用作从 O. formigenes 膜中纯化交换蛋白的指导。 蛋白质纯化后，实验将揭示动力学反应机制（乒乓反应或同时反应）以及反应循环中在膜上携带电荷的部分。 使用残基特异性化学探针、针对反向转运蛋白的单克隆抗体，以及衍生的氨基酸序列的物理结构预测。 产草酸杆菌是一种革兰氏阴性厌氧菌，其代谢能量的唯一来源来自草酸的脱羧作用。 已经确定，O. formigenes 的膜含有显示二价草酸盐和一价甲酸的生电交换的载体。 这种反应作为生物体维持质子动力的机制，对细胞生物化学特别感兴趣。 因此，前体和产物的反向输送导致负电荷的净进入，而草酸脱羧过程中质子的消耗改变了内部pH。 这是这种“辅助”载体的新用途，并且有利于确定利用羧酸的其他细胞是否也利用这种反向转运反应。 这项研究将检查这种载体的分子特性及其在细胞生物化学中的整体功能。 这项工作将有助于目前对细菌如何组织其细胞功能的理解。