LExEn: Genetic and Physiological Analysis of the Determinants of Halophillism in Halomonas Elongata

人们普遍认为地球上的生命起源于海水。虽然原始细胞是在Na+是最丰富的阳离子的环境中产生的，但生物已经进化到保持细胞质中的Na+浓度远远低于外部的Na+浓度。Na+在细胞膜上的不均匀分布所产生的势能被用来驱动不同的过程，如营养吸收、细胞运动和神经元的电活动。高浓度的氯化钠对大多数生物是有毒的，因此，高盐度是极端环境的一个例子。有少数生物体需要高浓度的氯化钠，这对大多数其他生物体都有抑制作用。依赖高浓度氯化钠的物种被称为嗜盐细菌。尽管人们普遍认为嗜盐微生物需要更高浓度的Na+来驱动营养吸收，但这一点尚未得到确凿的证明。最嗜盐的真细菌之一是长形盐单胞菌，它至少需要0.5M的氯化钠。这项研究项目正在验证这样的假设，即长臂藻需要高浓度的Na+，因为它的一些能量依赖的反应，如营养物质的运输或ATP的合成，是由Na+梯度驱动的。这种微生物特别适合分析嗜盐症的决定因素，因为它可以在简单定义的盐介质中生长，并且作为变形杆菌(包括大肠杆菌)g-分裂的一员，它可以进行遗传分析。本研究有以下具体目的：1)确定长盐单胞菌甘氨酸甜菜碱转运系统对Na+的需求量。之所以选择这种转运蛋白进行分析，是因为它已经在大肠杆菌中以功能形式表达，这使得测试它的活性是否与在长链杆菌的嗜盐背景中对Na+的要求类似。2)确定长臂藻中葡萄糖和氨基酸的运输系统是否对Na+有较高的需求。3)克隆长链球藻葡萄糖转运蛋白基因，并对该基因产物在长链球藻和大肠杆菌中对Na+的需求进行了比较研究。这些研究的实际好处是，它们将有助于开发长臂藻和其他嗜盐菌，用于在高盐度地区对环境污染物进行解毒。

It is generally accepted that life on Earth originated in sea water. Although primordial cells arose in an environment in which Na + is the most abundant cation, organisms have evolved to maintain the cytoplasmic concentration of Na + much lower than the external Na + concentration. The potential energy generated by unequal distribution of Na + across the cytoplasmic membrane is used to drive diverse processes, such as nutrient uptake, cell motility, and the electrical activity of neurons. NaCl at high concentrations is toxic to most organisms, and therefore, high salinity is one example of an extreme environment. There are a limited number of organisms which require high concentrations of NaCl that would be inhibitory to most other organisms. Species which are dependent on high concentrations of NaCl are called halophiles. Although it is generally believed that halophiles require elevated concentrations of Na + to drive nutrient uptake, this has not been proven conclusively. One of the most halophilic eubacteria is Halomonas elongata, which requires at least 0.5 M NaCl. The research project is testing the hypotheses that H. elongata requires high concentrations of Na + because some of its energy-dependent reactions, such as transport of nutrients or the synthesis of ATP, are driven by a Na + gradient. This organism is uniquely suitable for the analysis of the determinants of halophilism because it can grow in a simple defined salts medium and, as a member of the g-division of the Proteobacteria (which includes E. coli), it is amenable to genetic analysis. This research has the following specific aims: 1) Determine the Na+ requirement of the transport system for glycine betaine in Halomonas elongata. This transporter is chosen for analysis because it has already been expressed in functional form in E. coli, making it possible to test whether its activity has similar requirements for Na + in the non-halophilic background of E. coli as in the halophilic background of H. elongata. 2) Determine whether the transport systems for glucose and amino acids have high Na + requirement in H. elongata. 3) Clone the gene for the glucose transporter of H. elongata and carry out comparative studies of the Na + requirement of the product of this gene in H. elongata and in E. coli. These studies have the practical benefit that they will contribute to the development of H. elongata and other halophiles for the use of detoxification of environmental pollutants in highly saline areas.