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Specificity/Regulation of Cyanobacterial ABC Transporters

蓝藻 ABC 转运蛋白的特异性/调节

基本信息

批准号：
7263906
负责人：
Nicole M Koropatkin
金额：
$ 4.88万
依托单位：
DONALD DANFORTH PLANT SCIENCE CENTER
依托单位国家：
美国
项目类别：
财政年份：
2006
资助国家：
美国
起止时间：
2006-09-29 至 2008-09-28
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7263906
关键词：
ATP Hydrolysis ATP phosphohydrolase ATP-Binding Cassette Transporters ATPase Domain Affinity Bicarbonates Binding Binding Proteins Biochemical Biochemistry Biological Availability Carbon Carbon Dioxide Cell membrane Crystallography Cyanobacterium Cytoplasm Environment Enzymes Fellowship Genetic Individual Ions Iron Lead Ligand Binding Ligands Link Lipoprotein (a)Lipoprotein (a-)Lipoprotein Binding Membrane Metals Modeling Molecular Molecular Biology Techniques Names Nature Nitrates Nutrient Organism Oxygen Play Process Proteins Purpose Regulation Research Roentgen Rays Role Site-Directed Mutagenesis Specificity Structure Substrate Specificity Surface Plasmon Resonance Synechocystis Techniques Temperature Testing Thermodynamics Work Zinc base carbon fixation genetic manipulation greenhouse gases microcalorimetry mutant nitrate periplasm permease planetary Atmosphere protein protein interaction research study ribulose-1,5-bisphosphate solute uptake

项目摘要

DESCRIPTION (provided by applicant): The build-up of greenhouse gases in our atmosphere has, and will continue to, lead to a steady increase in the global temperature. A way to halt the further atmospheric accumulation of greenhouses gases, particularly carbon dioxide, is through manipulation of the natural carbon-sequestering mechanisms of aquatic cyanobacteria. One significant factor limiting carbon sequestration in these organisms is the low bioavailability of iron. For the acquisition of bicarbonate and iron, cyanobacteria employ high-affinity ATP-binding cassette (ABC) transporters. These ABC transporters are composed of a periplasmic solute-binding lipoprotein, a cytoplasmic-membrane spanning permease, and an ATPase that powers solute transport. A combination of x-ray crystallography, biochemical and molecular biology techniques will be used to examine both the molecular basis of substrate specificity and transport regulation in the bicarbonate CmpABCD, and the ferric iron FutABC transporters of Synechocystis sp PCC 6803. X-ray crystal structures of the CmpA, CmpC and CmpD proteins will be pursued, as well as a biochemical characterization of the unique multi-domain CmpC protein that is involved in the regulation of bicarbonate uptake. Both thermodynamic and enzymatic techniques will be employed to investigate the interactions between the ATPase domains of CmpC and CmpD. The FutABC transporter will be used to ask the question of whether the membrane permease and ATPase components of ABC transporters plays a role in solute recognition. The solute-binding specificity of the ferric-binding protein FutA1 will be altered in order to test whether the FutBC machinery can accept alternate metal ions. These experiments will involve genetic manipulation of Synechocystis for the purpose of testing metal ion acquisition through FutBC.

描述（由申请人提供）：大气中温室气体的积累已经并将继续导致全球气温稳步上升。阻止温室气体（特别是二氧化碳）在大气中进一步积累的一种方法是通过操纵水生蓝藻的天然固碳机制。限制这些生物体固碳的一个重要因素是铁的生物利用度低。为了获取碳酸氢盐和铁，蓝细菌采用高亲和力 ATP 结合盒 (ABC) 转运蛋白。这些 ABC 转运蛋白由周质溶质结合脂蛋白、跨细胞质膜通透酶和为溶质转运提供动力的 ATP 酶组成。将结合 X 射线晶体学、生化和分子生物学技术来检查碳酸氢盐 CmpABCD 和集胞藻 PCC 6803 的三价铁 FutABC 转运蛋白的底物特异性和转运调节的分子基础。将研究 CmpA、CmpC 和 CmpD 蛋白的 X 射线晶体结构，如下所示以及参与碳酸氢盐吸收调节的独特多结构域 CmpC 蛋白的生化特征。将采用热力学和酶技术来研究 CmpC 和 CmpD 的 ATPase 结构域之间的相互作用。 FutABC 转运蛋白将用于询问 ABC 转运蛋白的膜通透酶和 ATP 酶成分是否在溶质识别中发挥作用的问题。将改变铁结合蛋白 FutA1 的溶质结合特异性，以测试 FutBC 机器是否可以接受替代金属离子。这些实验将涉及集胞藻的基因操作，目的是测试通过 FutBC 获取金属离子。