Investigating Regulation of Integral Membrane Protein Cofactor Biosynthesis using

研究整合膜蛋白辅因子生物合成的调控

• 批准号: 8574527
• 负责人: Ronald F Peck
• 金额: $ 17.38万
• 依托单位: COLBY COLLEGE
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-03-01 至 2015-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8574527
• 关键词: Investigating; Regulation; Integral; Membrane; Protein

DESCRIPTION (provided by applicant): Integral membrane protein complexes consisting of proteins and small molecules that act as cofactors are ubiquitous and vitally important for organisms. For example, cytochrome oxidase is a mitochondrial complex that catalyzes the delivery of electrons to oxygen to allow cellular respiration. Rhodopsin is found in the eyes of many animals, including humans, and consists of an opsin membrane protein and a Vitamin A retinal cofactor. In these systems, an exquisite balance of protein and cofactor is maintained. Disruptions in this balance are thought to be causative factors for diseases such as Alzheimer's and retinitis pigmentosa. The proposed project describes a multidisciplinary approach to characterize a novel mechanism for coordinated synthesis of proteins and cofactors for integral membrane complexes. We will take advantage of the simple bacteriorhodopsin (BR) protein-cofactor complex in the microbe Halobacterium salinarum to potentially provide insight into how coordination is achieved in more complex systems. BR functions as a light-driven proton pump in the halophilic archaeon H. salinarum, and allows the organism to generate usable cellular energy under conditions where low oxygen prevents aerobic respiration. BR is the simplest possible membrane protein complex consisting of a single retinal cofactor bound to a single protein, bacterioopsin (BO). Under low oxygen conditions, H. salinarum rapidly induces BR biosynthesis, and this induction necessarily requires increased production of both the protein, BO, and the cofactor, retinal. In the proposed work, we explore how H. salinarum proteins interact to coordinate the synthesis of BO and retinal. Our preliminary results suggest that this coordination occurs by a novel mechanism where BO, not bound by retinal, inhibits an alternate biochemical pathway to only allow precursor molecules to be used for retinal synthesis. To investigate this mechanism, we will first use genetic and biochemical approaches to characterize enzymes that catalyze the synthesis of the retinal cofactor and related molecules. We will then identify which retinal precursors or enzymes interact with BO. Lastly, we will structurally characterize the interaction between BO and these molecules. PUBLIC HEALTH RELEVANCE: Molecular complexes consisting of proteins and cofactors our critical to our everyday life. For example, rhodopsin is a complex consisting of a protein, opsin, and a Vitamin A molecule that function together to allow us to see. Cytochrome oxidase is a complex of several proteins and a heme molecule that come together to allow us to get energy from the food we eat. Unfortunately, these cofactors and proteins can occasionally get out of balance leading to diseases such as retinitis pigmentosa and Alzheimer's. The proposed research will study a similar molecular complex in a microbe in order to elucidate the mechanisms that keep proteins and their cofactors in the correct balance.

描述（由申请人提供）：由蛋白质和作为辅因子的小分子组成的完整膜蛋白复合物无处不在，对生物体至关重要。例如，细胞色素氧化酶是一种线粒体复合体，它催化电子向氧气的传递，从而允许细胞呼吸。视紫红质存在于包括人类在内的许多动物的眼睛中，由视蛋白膜蛋白和维生素a视网膜辅助因子组成。在这些系统中，蛋白质和辅因子保持着微妙的平衡。这种平衡的破坏被认为是阿尔茨海默氏症和视网膜色素变性等疾病的致病因素。提议的项目描述了一个多学科的方法来表征一个新的机制协调合成的蛋白质和辅助因子的整体膜复合物。我们将利用盐盐杆菌中简单的细菌视紫红质（BR）蛋白-辅助因子复合体来潜在地洞察在更复杂的系统中如何实现协调。BR在嗜盐古菌H. salinarum中起着光驱动质子泵的作用，并允许生物体在低氧阻止有氧呼吸的条件下产生可用的细胞能量。BR是可能的最简单的膜蛋白复合物，由单一视网膜辅助因子与单一蛋白质细菌视素（BO）结合而成。在低氧条件下，H. salinarum迅速诱导BR的生物合成，而这种诱导必然需要增加蛋白质BO和辅助因子视网膜的产量。在这项工作中，我们探讨了盐芽胞杆菌蛋白如何相互作用以协调BO和视网膜的合成。我们的初步结果表明，这种协调是通过一种新机制发生的，其中BO不与视网膜结合，抑制另一种生化途径，仅允许前体分子用于视网膜合成。为了研究这一机制，我们将首先使用遗传和生化方法来表征催化视网膜辅助因子和相关分子合成的酶。然后我们将确定哪些视网膜前体或酶与BO相互作用。最后，我们将从结构上表征BO和这些分子之间的相互作用。