National Center for Quantitative Biology of Complex Systems

国家复杂系统定量生物学中心

• 批准号: 10089070
• 负责人: JOSHUA J COON
• 金额: $ 16.79万
• 依托单位: UNIVERSITY OF WISCONSIN-MADISON
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-07-05 至 2026-06-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10089070
• 关键词: Adherence; Amides; Amino Acids; Anions; Automobile Driving; Biology; Biomedical Research; Cations; Cells; Charge; Chromatography; Complex; Development; Dissociation; Electron Transport; Electrons; Fiber Optics; Free Radicals; Frequencies; Funding; Future; Generations; Glycopeptides; Glycosaminoglycans; Hybrids; Ions; Knowledge; Maps; Mass Spectrum Analysis; Methods; Modernization; Names; Nucleic Acids; Pathway interactions; Peptides; Photons; Post-Translational Protein Processing; Problem Solving; Process; Proteome; Proteomics; Reaction; Reagent; Recording of previous events; Research Personnel; Resources; Risk; Scanning; Speed; System; Technology; Time; Variant; Vendor; Vertebral column; base; density; design; experimental study; flexibility; glycosylation; improved; innovation; ion source; irradiation; mass analyzer; multiple omics; new technology; protein aminoacid sequence; protonation; reaction rate; small molecule; success; tandem mass spectrometry; user-friendly

PROJECT SUMMARY – TR&D 1 We direct this TR&D toward improved biomolecule characterization. The rationale is simple: you cannot quantify what you cannot characterize! Tandem mass spectrometry (MS/MS), the central approach for characterizing biomolecules with MS, is typically achieved by collisional activation. This process often fails, however, when the peptide precursor contains amino acids that inhibit random backbone protonation and PTMs that dissociate by a lower energy pathway than that involved in the cleavage of the amide linkage. Beyond these limitations, several biomolecule classes – O-glycopeptides, glycosaminoglycans, and modified nucleic acids, to name a few – preferentially ionize in the negative mode and are therefore extremely difficult to sequence with current technologies. To access these challenging biomolecules, researchers across the globe have turned to electron transfer dissociation (ETD). Now a mainstay in proteomic technology, ETD reacts peptide cations with small molecule anions. That said, future breakthroughs in biomolecule characterization will require key advancements in ETD technology. First, ETD reaction rates are now an order of magnitude longer (~60–90 ms per scan) than collision-based methods, limiting the impact ETD can have in a modern era that demands high throughput and high sensitivity. Second, the success rate of assigning peptide sequence to ETD scans suffers for precursors with low charge density (>1,000 m/z). That impedes ETD’s use in a number of applications, especially those involving large biomolecules or PTMs that add mass without adding charge (e.g., glycosylation). And finally, a variant of ETD – negative electron transfer dissociation (NETD) – has tremendous potential to allow access to acidic biomolecules, but is not widely available because there is not yet an ion source that can produce a high flux of the reagent cations needed to conduct the reaction. Here we leverage our deep history and expertise in ETD to propose creative, exportable technologies that remedy these crucial bottlenecks.

项目摘要 - TR＆D 1 我们将此TR＆D指向改善的生物分子表征。理由很简单：您无法量化 您无法表征的！串联质谱（MS/MS），这是表征表征的中心方法 具有MS的生物分子通常是通过碰撞激活来实现的。但是，当这个过程经常失败时，当 肽前体含有氨基酸，抑制随机主链质子化和PTM​​，而PTM被解离 比参与酰胺连接的裂解的能量途径较低。除了这些限制之外，有几个 生物分子类 - O-糖肽，糖胺聚糖和修饰的核酸，仅举几例 - 优先在负模式下电离，因此非常难以与电流进行测序 技术。为了获得这些挑战生物分子，全球的研究人员转向了电子 转移解离（ETD）。 ETD现在是蛋白质组学技术的中流tay柱，对小胡椒阳离子的反应小 分子阴离子。也就是说，生物分子表征的未来突破将需要关键的进步 在ETD技术中。首先，现在的ETD反应速率比数量级长（每次扫描约60–90毫秒） 基于碰撞的方法，限制了ETD在现代时代所产生的影响，该时代需要高吞吐量和 高灵敏度。第二，将肽序列分配给ETD扫描的成功率遭受了前体 低电荷密度（> 1,000 m/z）。这阻碍了ETD在许多应用中的使用，尤其是这些应用程序 涉及大型生物分子或PTM，它们增加质量而不会增加电荷（例如，糖基化）。最后，一个 ETD的变体 - 负电子传递解离（NETD）具有巨大的潜力，可以访问 酸性生物分子，但尚未广泛使用，因为尚无离子源可以产生高的离子源 进行反应所需的试剂阳离子的通量。在这里，我们利用了我们的深厚历史和专业知识 ETD提出创造性，可导出的技术，以记住这些关键的瓶颈。