Shared Resource Cores

共享资源核心

• 批准号: 8183701
• 负责人: GERALD Warren HART
• 金额: $ 62.86万
• 依托单位: JOHNS HOPKINS UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-07-01 至 2018-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8183701
• 关键词: Affinity; Amino Acids; Antibodies; Antibody Formation; Binding; Biological Assay; Body Fluids; COSY; Cardiac; Cardiovascular Diseases; Cell membrane; Cell surface; Cells; Chemicals; Chemistry; Collaborations; Complex; Consultations; Custom; Cytoplasm; Databases; Detection; Development; Discipline; Disease; Environment; Enzymes; Equipment; Glycoconjugates; Glycolipids; Glycopeptides; Glycoproteins; Goals; Grant; Hexosamines; In Vitro; Instruction; Isotope Labeling; Label; Laboratories; Lectin; Ligands; Link; Maps; Mass Spectrum Analysis; Membrane Proteins; Metabolic; Methodology; Methods; Mission; Modeling; Modification; Molecular Conformation; Monitor; NMR Spectroscopy; National Heart, Lung, and Blood Institute; Nucleotides; Oligosaccharides; Peptide antibodies; Peptides; Phosphorylated Peptide; Phosphorylation; Physiological; Polysaccharides; Post-Translational Protein Processing; Process; Property; Protein Analysis; Protein Glycosylation; Proteins; Proteomics; Protocols documentation; ROESY; Reagent; Regulation; Relative (related person); Research; Research Personnel; Research Project Grants; Resource Sharing; Resources; Role; Sampling; Screening procedure; Services; Site; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; Stable Isotope Labeling; Structure; TOCSY; Techniques; Technology; Tissues; Training; analog; analytical tool; base; design; extracellular; glycosylation; glycosyltransferase; human disease; in vivo; inhibitor/antagonist; interest; link protein; programs; protein complex; skills; small molecule; sugar; sugar nucleotide; tool; web site

PROJECT SU MMARY (See instructions): Analysis of glycoconjugate structure/functions requires specialized expertise from various disciplines. Many of these tools and approaches are not only difficult, but also require expensive and complex equipment. We have established our Shared Resource Core to not only contain the components that are most essential for the aims of this PEG, but also that build upon the unique strengths of our participating investigators. Each sub-Core is detailed in their separate sections. Sub-Core 01 - Mass Spectrometry -Drs. van Eyk and Zhang are leaders in mass spectrometric analysis of proteins and glycoproteins. The Core is extraordinarily well equipped. The Core's offerings include: 1) Identification of proteins; 2) global quantification of N-linked or O-linked glycoproteins using solidphase extraction of glycopeptides; 3) Lectin affinity/MS analyses; 4) Monitoring changes in glycosylation by lectin microarrays; 5) The use of "Click" chemistry for analyses of metabolically produced glycoconjugates; 6)Glycan profiling by MALDI-MS" (AXIMA-Resonance) and GlycanAnalyser. 7) Cell surface capture technology for isolation of glycoproteins; 2) Targeted capture of soluble glycoproteins from cytoplasm or extracellular environment; 3) MRM MS/MS assays for targeted quantification ofglycoproteins and glycosylation sites. Sub-Core C2 - NMR - When possible, glycan structure is best determined by NMR. Dr. Allen Bush is a leading expert in the structural analysis of glycans by NMR. His sub-Core supports the PEG by offering structural expertise for oligosaccharides, polysaccharides, glycopeptides, small glycoproteins and glycolipids. State of the art NMR equipment and methods are available in this sub-Core. Sub-Core C3 - Chemistry - Dr. Kevin Yarema is a leading chemical glycobiologist. His sub-Core will support the PEGs by: 1) providing sugar analogs for "metabolic glycoengineering" (eg. SCFA-hexosamines); 2) performing custom synthesis of inhibitors (eg. D-PDMP, TMG), azido sugars and sugar nucleotides, glycan-tagging tools and other small molecules. Sub-Core 04 - 0-GlcNAc Resources - Drs. Hart and Zachara are leaders in the analysis of 0-GlcNAc. In support of the PEGs we offer: 1) Tools for detecting 0-GlcNAc, OGT and OGA; 2) Develop site-specific 0-GlcNAc antibodies; 3) Provide synthetic 0-GlcNAc peptides for antibody production and MS standards; 4) Assays for OGT and OGA activity; 5) Quantitation of sugar nucleotides & nucleotides; 6) Assist with site mapping and quantification on single proteins or globally. This Shared Resource provides key technologies to allow rapid advancement of glycoscience research critical to the mission of the NHLBI.

项目摘要（参见说明）： 复合糖结构/功能的分析需要来自不同学科的专业知识。 其中许多工具和方法不仅困难，而且需要昂贵且复杂的设备。我们建立了共享资源核心，不仅包含实现该 PEG 目标最重要的组件，而且还建立在我们参与研究人员的独特优势之上。每个子核心在其单独的部分中都有详细介绍。 子核心 01 - 质谱 - 博士。 van Eyk 和Zhang 是蛋白质和糖蛋白质谱分析领域的领导者。核心装备非常精良。 Core 的产品包括： 1) 蛋白质鉴定； 2) 使用糖肽固相萃取对 N-连接或 O-连接糖蛋白进行整体定量； 3) 凝集素亲和力/MS分析； 4) 通过凝集素微阵列监测糖基化的变化； 5) 使用“点击”化学来分析代谢产生的糖复合物； 6) 通过 MALDI-MS" (AXIMA-Resonance) 和 GlycanAnalyser 进行聚糖分析。 7) 用于分离糖蛋白的细胞表面捕获技术；2) 从细胞质或细胞外环境中靶向捕获可溶性糖蛋白；3) 针对靶向的 MRM MS/MS 分析糖蛋白和糖基化位点的定量。 子核心 C2 - NMR - 如果可能，聚糖结构最好通过 NMR 确定。 Allen Bush 博士是利用 NMR 进行聚糖结构分析的领先专家。他的子核心通过提供寡糖、多糖、糖肽、小糖蛋白和糖脂的结构专业知识来支持 PEG。该子核心提供最先进的核磁共振设备和方法。 子核心 C3 - 化学 - Kevin Yarema 博士是一位领先的化学糖生物学家。他的子核心将通过以下方式支持 PEG：1）为“代谢糖工程”提供糖类似物（例如 SCFA-己糖胺）； 2) 进行抑制剂（例如D-PDMP、TMG）、叠氮糖和糖核苷酸、聚糖标签工具和其他小分子的定制合成。 子核心 04 - 0-GlcNAc 资源 - 博士。 Hart 和 Zachara 是 0-GlcNAc 分析领域的领导者。 为了支持 PEG，我们提供：1) 用于检测 0-GlcNAc、OGT 和 OGA 的工具； 2) 开发位点特异性0-GlcNAc抗体； 3) 提供合成的0-GlcNAc肽用于抗体生产和MS标准品； 4) OGT和OGA活性测定； 5) 糖核苷酸和核苷酸的定量； 6) 协助对单一蛋白质或全球蛋白质进行位点测绘和定量。该共享资源提供关键技术，以促进糖科学研究的快速发展，这对 NHLBI 的使命至关重要。