Synthesis of Glycogen in Bacteria and Starch in Plants: Evolution and Mechanism of Allosteric Control

细菌中糖原和植物中淀粉的合成：变构控制的进化和机制

• 批准号: 1616851
• 负责人: Miguel Ballicora
• 金额: $ 64万
• 依托单位: Loyola University of Chicago
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-08-01 至 2022-07-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1616851&HistoricalAwards=false
• 关键词: Synthesis; Glycogen; Bacteria; Starch; Plants

ADP-glucose pyrophosphorylase is central to starch synthesis in plants and glycogen synthesis in bacteria and other organisms, serving as carbon storage polymers. It is a common biotechnology target and an informed understanding of its regulation has implications for starch/glycogen synthesis in a host of organisms. Allosteric control of ADP-glucose pyrophosphorylase activity is an important but poorly understood regulatory mechanism. Presently it is unclear how the enzyme is allosterically activated and how that activation signal is transmitted structurally within the enzyme. The goal of this project is to perform computational, structural and biochemical studies to understand and characterize the allosteric signals and the interaction of the allosteric effectors on ADP-glucose pyrophosphorylase activity in bacteria as a model for plants. The PIs predict the presence of a common structural element that works as a switch but also the presence of an alternative allosteric signal binding site. This project will also contribute to education by training undergraduate and graduate students, establish international collaborations, and foster participation of underrepresented minorities in research by supporting programs such as McNair-SROP. This project will also support an international course on protein science that includes elements of the research, providing a unique training opportunity for undergraduate and graduate students. Plant starch and bacterial glycogen biosynthetic pathways provide models for the structure and evolution of allosteric regulatory functions of the enzyme ADP-glucose pyrophosphorylase. Despite the importance of these pathways, the allosteric mechanism that controls the activity of this enzyme is largely unknown. The project comprises computational, structural, and biochemical studies to understand and characterize the transmission of the allosteric signal and the interaction of the allosteric effectors in the ADP-glucose pyrophosphorylase family. It is predicted there is both a common structural element that works as a switch and an alternative allosteric signal guided by a secondary site. Three main research objectives will be pursued: 1) screen several enzyme forms from diverse branches of the phylogenetic tree for the presence of a dual allosteric mechanism; 2) obtain crystal structures of allosteric mutants ("switched off") in several different conditions with different ligands to test the hypothesis that certain residues form a conserved allosteric switch in ADP-glucose pyrophosphorylases; and 3) perform computational studies (molecular dynamics, correlated movement analysis, and network pathway analysis) to predict the signaling pathways of activation from each putative allosteric site.

ADP-葡萄糖焦磷酸化酶是植物中淀粉合成以及细菌和其他生物体中糖原合成的核心，充当碳储存聚合物。它是一个常见的生物技术目标，对其调控的深入了解对许多生物体中淀粉/糖原的合成具有影响。 ADP-葡萄糖焦磷酸化酶活性的变构控制是一种重要但知之甚少的调节机制。 目前尚不清楚酶如何变构激活以及激活信号如何在酶内结构上传递。该项目的目标是进行计算、结构和生化研究，以了解和表征变构信号以及变构效应子对细菌中 ADP-葡萄糖焦磷酸化酶活性的相互作用，作为植物模型。 PI 预测存在作为开关的通用结构元件，但也预测存在替代变构信号结合位点。该项目还将通过培训本科生和研究生、建立国际合作以及通过支持 McNair-SROP 等项目促进代表性不足的少数群体参与研究来促进教育。 该项目还将支持蛋白质科学国际课程，其中包括研究要素，为本科生和研究生提供独特的培训机会。 植物淀粉和细菌糖原生物合成途径为 ADP-葡萄糖焦磷酸化酶的变构调节功能的结构和进化提供了模型。尽管这些途径很重要，但控制这种酶活性的变构机制在很大程度上还是未知的。该项目包括计算、结构和生化研究，以了解和表征 ADP-葡萄糖焦磷酸化酶家族中变构信号的传递以及变构效应子的相互作用。 据预测，存在一个充当开关的共同结构元件和一个由辅助位点引导的替代变构信号。将追求三个主要研究目标：1）从系统发育树的不同分支中筛选几种酶形式，以确定是否存在双重变构机制； 2)在几种不同条件下用不同配体获得变构突变体（“关闭”）的晶体结构，以测试某些残基在ADP-葡萄糖焦磷酸化酶中形成保守变构开关的假设； 3) 进行计算研究（分子动力学、相关运动分析和网络通路分析）以预测每个假定的变构位点的激活信号通路。