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.

SU MMARY项目（请参阅说明）： 糖缀合物的结构/功能的分析需要各种学科的专业专业知识。 这些工具和方法中的许多不仅困难，而且需要昂贵且复杂的设备。我们已经建立了共享的资源核心，不仅包含对钉子目标最重要的组成部分，而且还基于参与调查人员的独特优势。每个子核在其单独的部分中详细介绍。 亚核01-质谱 - drs。 Van Eyk和Zhang是蛋白质和糖蛋白质谱分析的领导者。核心设备齐全。核心的产品包括：1）识别蛋白质； 2）使用糖肽的固相提取的N连锁或O连接糖蛋白的全球定量； 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缀合物； 6）MALDI-MS的聚糖分析（Axima-resonance）和糖纳拉塞菌。7）细胞表面捕获技术用于隔离糖蛋白； 2）针对细胞质或细胞外环境中可溶性糖蛋白的靶向捕获； 3）MMM/MS MMS/MS MMS/MS MMS/MS MMS/MS MMM/MS MMM/MS MMM/MS MMS/MS MMM/MSS分析。 亚核C2 -NMR-在可能的情况下，聚糖结构最好由NMR确定。艾伦·布什（Allen Bush）博士是NMR对聚糖结构分析的领先专家。他的子核通过提供寡糖，多糖，糖肽，小糖蛋白和糖脂的结构专业知识来支持PEG。该子核心有最先进的NMR设备和方法。 亚核C3-化学-Kevin Yarema博士是领先的化学糖生物学家。他的子核将通过：1）提供“代谢糖工程”的糖类似物（例如SCFA-HEXOSAMINE）； 2）执行抑制剂（例如D-PDMP，TMG），氮杂糖和糖核苷酸的自定义合成，聚糖引用工具和其他小分子。 子核04-0 -GLCNAC资源-DRS。 Hart和Zachara是0-GLCNAC分析的领导者。 为了支持PEG，我们提供的：1）用于检测0-GLCNAC，OGT和OGA的工具； 2）开发特异性的0-GLCNAC抗体； 3）为抗体产生和MS标准提供合成0-GLCNAC肽； 4）OGT和OGA活动的测定； 5）糖核苷酸和核苷酸的定量； 6）在单蛋白或全球范围内协助站点映射和定量。该共享资源提供了关键技术，以允许对NHLBI使命至关重要的糖基科学研究的快速发展。