Site-specific O-GlcNAc MAbs: New Tools for Glycoproteomics

位点特异性 O-GlcNAc MAb：糖蛋白组学的新工具

• 批准号: 8395145
• 负责人: RON ORLANDO
• 金额: $ 32.27万
• 依托单位: GLYCOSCIENTIFIC, LLC
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-09-17 至 2014-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8395145
• 关键词: Acetylglucosamine; Affinity; Antibodies; Antigens; Apoptotic; Autoantigens; Basic Science; Biological; Carbohydrates; Cell physiology; Complex; Cytoplasmic Protein; Detection; Diabetes Mellitus; Disease; Enzymes; Epitopes; Genetic Transcription; Gluconeogenesis; Glycopeptides; Harvest; Heart; Human; Hybridomas; Immune system; Immunization; Injectable; Insulin; Insulin Resistance; Lead; Letters; Libraries; Link; Modification; Monoclonal Antibodies; Mus; Nuclear Proteins; Phase; Phospho-Specific Antibodies; Phosphorylation; Play; Polysaccharides; Post-Translational Protein Processing; Preparation; Process; Production; Protein-Carbohydrate Interaction; Proteins; Reagent; Regulation; Research; Research Personnel; Resources; Role; Scientist; Screening procedure; Serine; Signal Transduction; Site; Specificity; Stimulus; Synthetic Vaccines; System; Testing; Threonine; Tumor Suppressor Genes; UDP-N-acetylglucosamine-peptide beta-N-acetylglucosaminyltransferase; Universities; Western Blotting; biological systems; candidate selection; cost; glycosylation; immunogenicity; medical schools; peptide O-linked N-acetylglucosamine-beta-N-acetylglucosaminidase; peptide structure; professor; protein aminoacid sequence; response; tool

DESCRIPTION (provided by applicant): O-glycosylation of serine and threonine of nuclear and cytoplasmic proteins by a single beta-N- acetyl-D-glucosamine moiety (O-GlcNAc) is a ubiquitous post-translational modification that is highly dynamic and fluctuates in response to cellular stimuli. O-GlcNAc often competes with protein phosphorylation, and these two modifications have extensive crosstalk in the regulation of signaling, transcription, and the functions of oncogenes and tumor suppressors. Diabetes, the biological driver for this application, is a prime example of a disease where increased levels of insulin resistance, inhibition of the anti-apoptotic action of insulin, alteration of circulating O-GlcNAc is known to disrupt normal signaling, and has been associated with the induction of adipocytokine levels, deregulation of gluconeogenesis, and modulation of insulin gene transcription. We will utilize a new immunogen strategy to develop site-specific O-GlcNac antibodies to four sites of O-GlcNAc modification on three proteins that play a role in signaling suppression in diabetes. Consequently, if we are successful, the mAbs generated in this initial study will have an immediate impact on diabetes research in addition to providing a test system for this strategy. We predict that at the end of phase 1, we will have demonstrated a strategy that will allow GlycoScientific to produce a wide range of O-GlcNAc site-specific MAbs, at a known and contained cost. We feel that these will have far reaching implications in disease research. PUBLIC HEALTH RELEVANCE: O-glycosylation of serine and threonine of nuclear and cytoplasmic proteins by a single beta-N- acetyl-D-glucosamine moiety (O-GlcNAc) is a ubiquitous post-translational modification that is highly dynamic and fluctuates in response to cellular stimuli. O-GlcNAc often competes with protein phosphorylation, and these two modifications have extensive crosstalk in the regulation of signaling, transcription, and the functions of oncogenes and tumor suppressors. Diabetes, the biological driver for this application, is a prime example of a disease where increased levels of insulin resistance, inhibition of the anti-apoptotic action of insulin, alteration of circulating O-GlcNAc is known to disrupt normal signaling, and has been associated with the induction of adipocytokine levels, deregulation of gluconeogenesis, and modulation of insulin gene transcription. We will utilize a new immunogen strategy to develop site-specific O-GlcNac antibodies to four sites of O-GlcNAc modification on three proteins that play a role in signaling suppression in diabetes. Consequently, if we are successful, the mAbs generated in this initial study will have an immediate impact on diabetes research in addition to providing a test system for this strategy.

描述（由申请人提供）：单个β-N-乙酰基-D-葡萄糖胺部分（O-GlcNAc）对核蛋白和细胞质蛋白的丝氨酸和苏氨酸进行O-糖基化是一种普遍存在的翻译后修饰，具有高度动态性，并随细胞刺激而波动。O-GlcNAc经常与蛋白质磷酸化竞争，并且这两种修饰在信号传导、转录以及癌基因和肿瘤抑制因子的功能的调节中具有广泛的串扰。糖尿病是本申请的生物学驱动因素，是已知胰岛素抗性水平增加、胰岛素抗凋亡作用抑制、循环O-GlcNAc改变破坏正常信号传导的疾病的主要实例，并且与脂肪细胞因子水平诱导、脂肪生成失调和胰岛素基因转录调节相关。 我们将利用一种新的免疫原策略来开发针对三种蛋白质上的O-GlcNAc修饰的四个位点的位点特异性O-GlcNAc抗体，这些蛋白质在糖尿病的信号传导抑制中发挥作用。因此，如果我们成功了，在这项初步研究中产生的单克隆抗体将对糖尿病研究产生直接影响，并为这一策略提供测试系统。 我们预测，在第1阶段结束时，我们将展示一种策略，该策略将允许GlycoScientific以已知和受控的成本生产广泛的O-GlcNAc位点特异性单克隆抗体。我们认为这些将对疾病研究产生深远的影响。 公共卫生相关性：通过单个β-N-乙酰基-D-葡糖胺部分（O-GlcNAc）对细胞核和细胞质蛋白的丝氨酸和苏氨酸的O-糖基化是普遍存在的翻译后修饰，其是高度动态的并且响应于细胞刺激而波动。O-GlcNAc经常与蛋白质磷酸化竞争，并且这两种修饰在信号传导、转录以及癌基因和肿瘤抑制因子的功能的调节中具有广泛的串扰。糖尿病是本申请的生物学驱动因素，是已知胰岛素抗性水平增加、胰岛素抗凋亡作用抑制、循环O-GlcNAc改变破坏正常信号传导的疾病的主要实例，并且与脂肪细胞因子水平诱导、脂肪生成失调和胰岛素基因转录调节相关。 我们将利用一种新的免疫原策略来开发针对三种蛋白质上的O-GlcNAc修饰的四个位点的位点特异性O-GlcNAc抗体，这些蛋白质在糖尿病的信号传导抑制中发挥作用。因此，如果我们成功了，在这项初步研究中产生的单克隆抗体将对糖尿病研究产生直接影响，并为这一策略提供测试系统。