Post-Translation Processing of Alpha-Dystroglycan

α-肌营养不良聚糖的翻译后加工

• 批准号: 7691724
• 负责人: DAVID H LIVE
• 金额: $ 16.23万
• 依托单位: UNIVERSITY OF GEORGIA
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-09-22 至 2011-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7691724
• 关键词: Acetylglucosamine; Address; Biochemical; Biochemical Reaction; Biological Assay; Carbohydrates; Clinical; Complement; Complex; Defect; Detection; Development; Disease; Dystroglycan; Dystrophin; Enzymes; Evaluation; Extracellular Matrix; Glycopeptides; Glycoproteins; Goals; Inherited; Intervention; Length; Link; Mannose; Maps; Mass Spectrum Analysis; Methods; Modification; Molecular; Mucins; Muscle Cells; Muscle eye brain disease; Muscular Dystrophies; Pattern; Physiological; Polypeptide N-acetylgalactosaminyltransferase; Polysaccharides; Post-Translational Protein Processing; Process; Property; Proteins; Research; Role; Series; Site; Sorting - Cell Movement; Specificity; Staging; Structure; Structure-Activity Relationship; Therapeutic; Therapeutic Intervention; Tissues; Touch sensation; Translation Process; Up-Regulation; Vertebral column; Vision; alpha Dystroglycan; base; chemical synthesis; congenital muscular dystrophy; design; dystroglycan 1; enzyme activity; enzyme substrate; gene therapy; glycosylation; glycosyltransferase; improved; insight; interest; intermolecular interaction; novel; polypeptide; protein O-mannose beta-1,2-N-acetylglucosaminyltransferase; public health relevance; therapy design; tool

DESCRIPTION (provided by applicant): The objective of the proposed research is to understand the post-translational processing of 1- dystroglycan (1-DG), a glycoprotein that is a key component of the dystrophin-glycoprotein complex, anchoring muscle cells to the extracellular matrix. Defective glycosylation of 1-DG in its highly conserved central mucin-like region, with numerous glycosylated S and T residues is the cause of several forms of hereditary muscular dystrophy. Pendant glycans are linked to the protein by either N-acetylglucosamine (GalNAc) or the more unusual mannose (Man) residue whose sites are now just being mapped. Relationships between problems in assembly of the O-Man tetra-saccharide and disease have been established. We have recently found evidence that the GalNAc residues are important structurally. Synthetic, biochemical and structural methods will be integrated in the research to develop an understanding of the steps in this complex process, providing a quantitative description of enzymatic transformations and also facilitating insights into key interactions in which 1-DG participates. An understanding of the steps in this process will provide basis for rational design of therapies to correct the defects, including gene therapy and up-regulation of other enzymes that may complement the defective ones. The focus will be on two steps following the initial O-Man modifications of 1-DG. The two major aims are 1) to analyze the substrate profile of POMGnT1, an enzyme involved in a key step in the O-Man glycan assembly whose defects are associated with muscle-eye-brain disease, to better understand how rescue the defect and to develop a better assay for detection of defective enzyme, and 2) to elucidate the activity of polypeptide GalNAc transferases in initiating O-GalNAc sites on 1-DG and the relationship of these to POMGnT1 glycosylation. PUBLIC HEALTH RELEVANCE: Several forms of muscular dystrophy are associated with aberrations in the attachment of carbohydrates to the glycoprotein 1-dystroglycan arising from defects in the enzymes that carry out the complex series of steps leading to the specific installation of carbohydrates on it. The proposed research would use an integrated approach combining chemical synthesis of glycopeptides from1- dystroglycan, biochemical analysis of their modification by glycosyltransferase enzymes, and structural analysis of the reactants and products, to develop an understanding at a molecular level of the post-translational modification of this important glycoprotein. The information derived will contribute to understanding the pathological mechanisms of several forms of muscular dystrophies, improving clinical assay, and rational design of therapeutic interventions.

描述（由申请人提供）：拟议研究的目的是了解1-肌营养不良聚糖（1- dg）的翻译后加工，1- dg是一种糖蛋白，是肌营养不良蛋白-糖蛋白复合物的关键成分，将肌肉细胞锚定在细胞外基质上。1-DG高度保守的中心黏液样区域的糖基化缺陷，以及大量糖基化的S和T残基，是几种形式的遗传性肌营养不良的原因。悬垂聚糖通过n -乙酰氨基葡萄糖（GalNAc）或更不寻常的甘露糖（Man）残基与蛋白质相连，其位置现在刚刚被绘制出来。已经建立了O-Man四糖组装问题与疾病之间的关系。我们最近发现的证据表明GalNAc残基在结构上是重要的。合成、生化和结构方法将整合到研究中，以了解这一复杂过程中的步骤，提供酶转化的定量描述，并促进对1-DG参与的关键相互作用的见解。了解这一过程中的步骤将为合理设计纠正缺陷的治疗方法提供基础，包括基因治疗和上调其他可能补充缺陷酶的酶。重点将放在1-DG初始O-Man修改后的两个步骤上。研究的两个主要目的是：1)分析POMGnT1（一种参与O-Man聚糖组装关键步骤的酶，其缺陷与肌肉-眼-脑疾病相关）的底物谱，以更好地了解如何修复缺陷并开发更好的检测缺陷酶的方法；2)阐明多肽GalNAc转移酶在启动1- dg上O-GalNAc位点的活性及其与POMGnT1糖基化的关系。公共卫生相关性：几种形式的肌营养不良与碳水化合物附着在糖蛋白1-三聚糖聚糖上的畸变有关，这种畸变是由执行导致碳水化合物在其上特定安装的一系列复杂步骤的酶的缺陷引起的。本研究将采用综合的方法，结合1-三聚糖聚糖的糖肽化学合成，糖基转移酶修饰的生化分析，以及反应物和产物的结构分析，在分子水平上了解这种重要糖蛋白的翻译后修饰。所得的信息将有助于理解几种形式的肌肉萎缩症的病理机制，改进临床检测，合理设计治疗干预措施。