Posttranslational Regulation of Pep-Carboxylase Activity in Higher Plants

高等植物 Pep 羧化酶活性的翻译后调控

• 批准号: 9315928
• 负责人: Raymond Chollet
• 金额: $ 42.4万
• 依托单位: University of Nebraska-Lincoln
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 1994
• 资助国家: 美国
• 起止时间: 1994-05-01 至 1998-04-30
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=9315928&HistoricalAwards=false
• 关键词: Posttranslational; Regulation; Pep; Carboxylase; Activity

9315928 Chollet The experimental plan is composed of three areas of research on the control of higher plant phosphoenolpyruvate carboxylase (PEPC) activity in vivo by regulatory phosphorylation and allosteric metabolite effectors. Specific research objectives include (i) the continued scrutiny of the highly regulated PEPC-kinase signal- transduction chain by in situ studies with intact C4 and CAM plants using an integrated biochemical, immunological and/or molecular- genetic approach, and the further elucidation of the complex C4 PEPC-kinase signal-transduction chain by in situ studies with intact C4-mesophyll cells and protoplasts. The outcome of these objectives could strongly affect our basic understanding of the regulation of PEPC activity by L-malate, the negative allosteric effector. Conversely, project (ii) will focus on the opposing control of C4/CAM PEPC activity exerted by positive phosphorylated effectors (e.g., glucose 6-P, triose-P) by mapping this activator binding domain with photoaffinity-labeling and site-directed mutagenesis techniques. Finally, project (iii) will critically assess the possible regulatory phosphorylation of nonphotosynthetic PEPC in leaves and root-nodules from C3 plants by integrative in vivo (detached leaves and nodules), in situ (intact leaf protoplasts), and in vitro (isolated PEPC and PEPC-kinase) analysis. Results from these three projects will not only delineate important details of the control of C4- and CAM- photosynthesis, and details of general C/N metabolism in plants at the level of cytosolic PEPC, but at the same time will provide much needed insight into enzyme regulation in higher plants by reversible protein phosphorylation. %%% This project is focused on the posttranslational regulation of the carbon dioxide (CO2)-fixing enzyme phosphoenolpyruvate carboxylase (PEPC) in higher plants. While this enzyme is best known for its role in CO2 fixation during C4 photosynthesis and Crassulacean acid metabolism (CA M) in crops as maize, sorghum, sugarcane, and pineapple (CAM), it also functions in general plant carbon/nitrogen metabolism. Besides these important physiological and functional considerations, the PEPC enzyme represents one of the few well- known examples in higher plants of an enzyme that is controlled by the opposing action of a protein phosphatase and a protein-serine kinase. Consequently, this complex regulatory phosphorylation cycle forms the central focus of many of our present studies. Understanding about how the cycle operates is important in regulation of CO2 assimilation and in plant metabolism in general. It can form the basis for biotechnological engineering of plants to improve growth and productivity. This project is supported jointly by Metabolic Biochemistry Program, MCB and the Plant Integrative Biology Program, IBN. ***

9315928 Chollet 该实验计划由三个研究领域组成，通过调节磷酸化和变构代谢效应物控制高等植物体内磷酸烯醇丙酮酸羧化酶（PEPC）活性。 具体研究目标包括 (i) 通过使用综合生化、免疫学和/或分子遗传学方法对完整 C4 和 CAM 植物进行原位研究，持续审查高度调控的 PEPC 激酶信号转导链，并通过对完整 C4 叶肉细胞和原生质体进行原位研究，进一步阐明复杂的 C4 PEPC 激酶信号转导链。 这些目标的结果可能会强烈影响我们对负变构效应物 L-苹果酸调节 PEPC 活性的基本理解。 相反，项目 (ii) 将重点关注正磷酸化效​​应子（例如葡萄糖 6-P、丙糖-P）对 C4/CAM PEPC 活性的反向控制，方法是使用光亲和标记和定点诱变技术绘制该激活剂结合结构域。 最后，项目 (iii) 将通过体内（分离的叶子和根瘤）、原位（完整的叶子原生质体）和体外（分离的 PEPC 和 PEPC 激酶）综合分析，严格评估 C3 植物叶子和根瘤中非光合 PEPC 可能的调节磷酸化。 这三个项目的结果不仅将描述 C4 和 CAM 光合作用控制的重要细节，以及植物中细胞质 PEPC 水平上一般 C/N 代谢的细节，同时还将提供对高等植物中通过可逆蛋白质磷酸化进行酶调节的急需的见解。 %%% 该项目的重点是高等植物中二氧化碳 (CO2) 固定酶磷酸烯醇丙酮酸羧化酶 (PEPC) 的翻译后调控。 虽然这种酶以其在玉米、高粱、甘蔗和菠萝 (CAM) 等作物的 C4 光合作用和景天酸代谢 (CAM) 过程中的 CO2 固定作用而闻名，但它也在一般植物碳/氮代谢中发挥作用。 除了这些重要的生理和功能考虑因素之外，PEPC 酶代表了高等植物中少数众所周知的酶实例之一，该酶由蛋白磷酸酶和蛋白丝氨酸激酶的相反作用控制。 因此，这种复杂的调节磷酸化循环构成了我们目前许多研究的中心焦点。 了解循环的运作方式对于调节二氧化碳同化和植物代谢非常重要。 它可以构成植物生物技术工程的基础，以提高植物的生长和生产力。 该项目由 MCB 代谢生物化学计划和 IBN 植物综合生物学计划联合支持。 ***