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REGULATION OF GENE EXPRESSION IN HALOBACTERIUM HALOBIUM

卤杆菌基因表达的调控

基本信息

批准号：
3300506
负责人：
SHILADITYA DASSARMA
金额：
$ 12.17万
依托单位：
UNIVERSITY OF MASSACHUSETTS AMHERST
依托单位国家：
美国
项目类别：
财政年份：
1989
资助国家：
美国
起止时间：
1989-04-01 至 1994-03-31
项目状态：
已结题

项目摘要

Halobacterium halobium is a very unusual microorganism that grows optimally in nearly saturated brine. Although H. halobium usually generates energy for growth by oxidative phosphorylation, it has the capacity for anaerobic growth by photophosphorylation. For photophosphorylation, the light-driven proton pumping activity of the purple membrane protein, bacteriorhodopsin, is necessary. Phototrophic growth also requires medium supplemented with retinal, the purple membrane chromophore, biosynthesis of which requires oxygen. H. halobium induces the synthesis of bacteriorhodopsin and also the intracellular organelles called gas vesicles when grown under limiting oxygen and high light intensity. Gas vesicles allow cells to float to the surface of the culture, thus, increasing the availability of oxygen and light. We have recently determined that the induction of both purple membrane and gas vesicles occurs at the transcriptional level. Thus, we have in hand a very interesting system for studying the molecular mechanisms of genetic regulation in H. halobium. We already have the necessary tools to carry out the proposed study, including cloned purple membrane (bop) and gas vesicle (gvp) genes, regulatory mutants, and assays for both gas vesicle and purple membrane content and mRNA levels. The following studies are planned: 1) We will re-examine the parameters that regulate gas vesicle and purple membrane synthesis, including the role of oxygen and light as well as other environmental and nutritional parameters. 2) We will also investigate the mechanism of coordinate synthesis of the purple membrane opsin and chromophore. 3) other H. halobium genes exhibiting similar modes of regulation will be isolated by differential hybridization techniques. 4) The promoter-regulatory region of bop, gvp and other coordinately regulated genes will be examined in the wild-type and regulatory mutants by in vivo footprinting experiments under various growth conditions. 5) Selectable shuttle vectors will be constructed for transfer of genes between H. halobium and E. coli. Important long-term goals would be to understand the precise biophysical mechanisms of gene regulation operating in the extremely saline cytoplasm of H. halobium. Knowledge about genetic regulation is important to health for many reasons such as the fact that many diseases result from mutation of normal regulatory properties.

盐生盐杆菌是一种非常不寻常的微生物，最好在接近饱和的盐水中。虽然H.盐生菌通常通过氧化磷酸化为生长提供能量，厌氧生长的能力，通过磷酸化。为磷酸化，光驱动的质子泵活性，紫膜蛋白细菌视紫红质是必需的。光养生长还需要补充有视黄醛的培养基，紫膜发色团，其生物合成需要氧气 H.盐诱导细菌视紫红质的合成，细胞内的细胞器也叫气泡，在有限的氧气和高光强下。气体囊泡允许细胞漂浮到培养物的表面，因此，增加了细胞的增殖。氧气和光的可用性。我们最近确定，紫膜和气泡的诱导都发生在转录水平。因此，我们手中有一个非常一个有趣的系统，用于研究遗传的分子机制，调节H.盐生菌。我们已经有了必要的工具，开展拟议的研究，包括克隆紫膜 (bop)和气体囊泡（GVP）基因、调节突变体和测定对气泡和紫膜含量及mRNA水平的影响。计划进行以下研究：1）我们将重新研究调节气泡和紫膜合成的参数，包括氧气和光以及其他的作用环境和营养参数。 2)我们还将探讨了紫色配合物的合成机理膜视蛋白和发色团。 3)其他H.盐生菌基因表现出类似的调节模式将被隔离，差异杂交技术。 4)发起人-监管者 bop、gvp等协同调控基因的区域，在野生型和调节突变体中通过体内各种生长条件下的足迹实验。第五章）将构建选择性穿梭载体用于转移 H. halobium和E.杆菌重要的长期目标是理解基因的精确生物物理机制调节在H. 盐生菌。关于基因调控的知识对于健康的原因很多，例如许多疾病导致正常调节特性的突变。