Initiation and regulation of mucin-type O-glycosylation

粘蛋白型O-糖基化的启动和调节

基本信息

批准号：
9012950
负责人：
THOMAS A GERKEN
金额：
$ 15.74万
依托单位：
CASE WESTERN RESERVE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2015
资助国家：
美国
起止时间：
2015-01-01 至 2018-11-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9012950
关键词：
Accounting Address Animal Model Animals Belief Benign Biological Biological Process Biology C-terminal Cardiovascular Diseases Cardiovascular system Cell Communication Cell Culture Techniques Cell Line Cells Complement Data Development Disease Embryonic Development Enzymatic Biochemistry Enzymes Family Fertility Functional disorder Glycopeptides Goals Health Hormonal Host Defense Human In Vitro Individual Inflammatory Institution Kinetics Knock-out Lead Learning Libraries Link Malignant Neoplasms Membrane Metabolic Methods Modification Molecular Mucins Mus Mutation Online Systems Pattern Peptides Play Polypeptide N-acetylgalactosaminyltransferase Polysaccharides Prevalence Process Property Protein Glycosylation Protein Isoforms Proteins Regulation Reporting Reproduction Role Series Site Specificity Structure Substrate Specificity Therapeutic Tissues Transferase Work animal tissue base design fly glycosylation in vivo inhibitor/antagonist innovation male man member neoplastic novel novel strategies polypeptide ppGalNAc-T predictive modeling preference public health relevance sugar therapeutic target tool

项目摘要

DESCRIPTION (provided by applicant): Most membrane and secreted proteins are decorated with mucin-type O-glycans which serve diverse biological roles. The mechanisms behind their biological roles and more specifically what governs O-glycan site selection and subsequent O-glycan elongation are largely unknown. The objective of this project is to elucidate the processes governing O-glycan site selection and O-glycan elongation at the enzyme and peptide substrate level in order to address the molecular mechanisms and biology of O-glycosylation. Mucin type O- glycosylation is vital to animal reproduction and development and increasingly linked to a wide range of rare to common disease states (such as hormonal/metabolic dysfunction, impaired host defense, inflammatory and cardiovascular diseases and even cancers) where it serves to modulate diverse biological functions including cell-cell interactions. Many disorders are linked to changes in expression or mutation of individual members of the large family (20 in man) of polypeptide-GalNAc transferases (ppGalNAc Ts) that initiate O-glycosylation by adding GalNAc to polypeptide Ser or Thr residues. O-glycosylation is absolutely required for embryonic development of the mouse and fly, relying on the Core 1 elongating transferase (T-synthase) in the mouse and several individual ppGalNAc T isoforms in the fly. How individual ppGalNAc T isoforms in this large family (or even T-synthase) can play such critical biological roles is unknown as the substrate specificity of these transferases have not been sufficiently characterized. That ppGalNAc T site selection is also modulated (positively or negatively) by prior glycosylation underscores its complexity and how much more needs to be learned of its specificity. The detailed characterization of ppGalNAc T isoform and elongating transferase specificity proposed in this project will lead to our understanding of the molecular mechanisms underlying the biological roles of O-glycosylation and will eventually lead to novel strategies to treat diseases of aberrant O- glycosylation. The AIMS of this project are to (1) expand the innovative use of a library of novel random peptide and glycopeptide substrates to fully characterize the specificity and basic enzymology of the transferases that initiate and elongate mucin type O-glycosylation, (2) to use these data to further develop sophisticated web based O-glycan predictive tools that include ppGalNAc T isoform and elongating transferase peptide and glycopeptide specificity, and (3) to develop a novel method for identifying in vivo isoform specific glycosylation targets using tissues from transferase knock-out animal models. These basic studies together with those of colleagues at other institutions will advance our understanding of the properties of these transferases, their targets (and resultant glycan structures) and ultimately the mechanisms of their biological role and function. These studies will significantly advance the field and will allw the development of novel specific inhibitors for potential use as targeted therapeutics.

描述（由申请人提供）：大多数膜和分泌蛋白都装饰有粘蛋白型 O-聚糖，其具有多种生物学作用，其生物学作用背后的机制，更具体地说，是控制 O-聚糖位点选择和随后的 O-聚糖延伸的机制。该项目的目的是阐明在酶和肽底物水平上控制 O-聚糖位点选择和 O-聚糖延伸的过程，以解决分子机制和生物学问题。粘蛋白型 O-糖基化对于动物的繁殖和发育至关重要，并且与各种罕见到常见的疾病状态（例如激素/代谢功能障碍、宿主防御受损、炎症和心血管疾病甚至癌症）密切相关。它可以调节多种生物功能，包括细胞间相互作用，许多疾病与多肽-GalNAc 转移酶 (ppGalNAc) 大家族（人类有 20 个）的表达或突变有关。通过向多肽 Ser 或 Thr 残基添加 GalNAc 来启动 O-糖基化的 Ts) 对于小鼠和果蝇的胚胎发育来说是绝对必需的，这依赖于小鼠和一些个体中的 Core 1 延伸转移酶（T-合酶）。果蝇中的 ppGalNAc T 同工型在这个大家族（甚至 T 合酶）中作为底物如何发挥如此重要的生物学作用尚不清楚。这些转移酶的特异性尚未得到充分表征，ppGalNAc T 位点选择也受到先前糖基化的调节（正向或负向），这强调了其复杂性以及需要了解其特异性的更多信息。该项目中提出的转移酶特异性将有助于我们了解 O-糖基化生物学作用背后的分子机制，并最终导致治疗异常疾病的新策略。该项目的目标是 (1) 扩展新型随机肽和糖肽底物库的创新用途，以充分表征启动和延长粘蛋白型 O-糖基化的转移酶的特异性和基本酶学，（ 2) 使用这些数据进一步开发基于网络的复杂 O-聚糖预测工具，其中包括 ppGalNAc T 同工型以及延长转移酶肽和糖肽特异性，以及(3) 开发一种利用组织识别体内异构体特异性糖基化靶标的新方法这些基础研究与其他机构的同事的研究将促进我们对这些转移酶的特性、它们的靶标（以及由此产生的聚糖结构）以及最终它们的生物学作用和功能研究的机制的理解。将显着推进该领域的发展，并将允许开发新型特异性抑制剂，以用作靶向治疗。