LExEN: The Chemistry and Biology of Unusual Adaptive Solutes in Archaeoglobus Fulgidus and Natronococcus Occultus

LExEN：Archaeoglobus Fulgidus 和 Natronococcus Occultus 中异常适应性溶质的化学和生物学

• 批准号: 9978250
• 负责人: Mary Roberts
• 金额: $ 37.21万
• 依托单位: Boston College
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-09-01 至 2003-10-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=9978250&HistoricalAwards=false
• 关键词: LExEN; Chemistry; Biology; Unusual; Adaptive

Roberts9978250Two archaea, Archaeoglobus fulgidus and Natronococcus occultus, grow in extreme environments. Although both of these grow in moderate to high salt media (with N. occultus a true halophile), A. fulgidus is a hyperthermophile while N. occultus requires an external pH of 9.5 and is classified as a haloalkaliphile.It has been proposed that organisms that live and adapt to hypersaline environments can provide insightsinto life on early Mars. Halophilic archaea, in particular, may be important models for life on early Marswith sulfate-reducing archaea providing the most realistic models for extra-terrestrial environments. This project seeks to understand how both A. fulgidus and N. occultus respond to changes in external NaCl, including protein and solute (organic osmolyte) synthesis, K+ fluxes, and mechanisms for stabilization of intracellular macromolecules. The emphasis is on the biology of several unusual chemical compounds used as organic osmolytes, and their very specific role in the survival of these cells. Techniques used include in vivo NMR spectroscopy, atomic absorption, 35S-uptake and 2D SDS-PAGE, and computer simulations of water and the effect osmolytes have on water structure and dynamics. For A. fulgidus an important area to investigate is how growth temperature regulates di-inositol-1,1'-phosphate (DIP) synthesis and accumulation. This unusual organic solute is only accumulated and used to balance external osmotic pressure when the organism is grown above 80 degrees C. Cloning, overexpression, and characterization of enzymes involved in DIP synthesis will provide details on the biosynthetic machinery for this solute whose accumulation correlates with thermophily in many other archaea as well. Where possible overexpressed enzymes will be crystallized for structure determination. This goal will be particularly important for the novel A. fulgidus inositol-1-phosphate synthase that appears to be a class II aldolase, and for the proposed CTP:I-1-P cytidylyltransferase and DIP synthase activities (when they are identified and purified from the cells). Monitoring protein and message synthesis for these enzymes upon thermal stress should provide further insights into the temperature regulation of DIP accumulation. A related phosphodiester, di-a-glycerol phosphate, appears to be the major osmolyte at lower growth temperatures in A. fulgidus. Nothing is known about the biosynthesis of this solute from a-glycerol phosphate (although it is likely an activated glycerol phosphate is condensed with glycerol in analogy to the 'DIP synthase' reaction). NMR experiments are planned to elucidate the enzymology involved.

两种古细菌，闪烁古生菌和隐钠球菌，生长在极端的环境中。 虽然这两种植物都生长在中等至高盐的培养基中（N。occultus a true halophile），A. fulgidus为超嗜热菌，N. occultus需要外部pH值为9.5，并被归类为一个haloalkaliphile。有人提出，生活和适应高盐环境的有机体可以提供洞察早期火星上的生命。特别是嗜盐古生菌，可能是早期火星生命的重要模型，硫酸盐还原古生菌为地外环境提供了最真实的模型。这个项目旨在了解如何既A。fulgidus和N. occultus响应外部NaCl的变化，包括蛋白质和溶质（有机渗透压）的合成，K+流量，和细胞内大分子的稳定机制。重点是几种不寻常的化合物作为有机渗透剂的生物学，以及它们在这些细胞生存中的特殊作用。所使用的技术包括体内NMR光谱，原子吸收，35 S-摄取和2D SDS-PAGE，以及水的计算机模拟和渗透剂对水结构和动力学的影响。 对于.研究的一个重要领域是生长温度如何调节二肌醇-1，1 '-磷酸（DIP）的合成和积累。这种不寻常的有机溶质只有在生物体生长在80摄氏度以上时才会积累并用于平衡外部渗透压。 克隆，过表达，和DIP合成所涉及的酶的表征将提供详细的生物合成机制，这种溶质的积累与嗜热性在许多其他古菌以及。在可能的情况下，将过表达的酶结晶用于结构测定。这一目标对于小说A来说尤其重要。闪电肌醇-1-磷酸合酶，似乎是II类醛缩酶，并具有拟议的CTP：I-1-P胞苷酰转移酶和DIP合酶活性（当它们从细胞中鉴定和纯化时）。监测蛋白质和信息的合成，这些酶在热应力应提供进一步的见解DIP积累的温度调节。一个相关的磷酸二酯，二-α-甘油磷酸，似乎是主要的渗透剂在较低的生长温度在A。闪电。关于这种溶质从α-甘油磷酸的生物合成是未知的（尽管类似于“DIP合成酶”反应，活化的甘油磷酸可能与甘油缩合）。 核磁共振实验计划阐明所涉及的酶学。