Assessing the Biomolecular Structures that Result from Electrospray Ionization

评估电喷雾电离产生的生物分子结构

• 批准号: 10712440
• 负责人: Elyssia S Gallagher
• 金额: $ 33.64万
• 依托单位: BAYLOR UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-09-01 至 2028-08-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10712440
• 关键词: Address; Affect; Binding; Biological; Biological Process; Carbohydrates; Cells; Cellular biology; Charge; Complex; Development; Disease; Disease Progression; Electrospray Ionization; Exhibits; Future; Gases; Glycoconjugates; Goals; Infection; Ions; Isomerism; Ligand Binding; Mass Spectrum Analysis; Metals; Methodology; Methods; Modeling; Molecular; Phase; Polymers; Polysaccharides; Process; Property; Proteins; Research; Research Personnel; Signal Transduction; Sodium; Spectrometry, Mass, Electrospray Ionization; Structure; Techniques; Validation; Work; adduct; analytical method; deprotonation; host-microbe interactions; improved; insight; ion mobility; ionization; ionization technique; molecular dynamics; molecular recognition; novel; pathogen; protein complex; protein structure; protonation; tandem mass spectrometry; tool

Project Summary / Abstract Glycans, heterogeneous polymers of carbohydrates, interact with proteins to initiate a multitude of biologi- cal processes, including molecular recognition, cellular signaling, and host-microbe interactions. Mass spec- trometry (MS) methods have become powerful tools for characterizing the structures of glycans and their inter- actions with proteins. Electrospray ionization (ESI) is a common ionization method for transferring these bio- molecules from solution to the gas-phase for MS analysis. However, both glycans and proteins exhibit struc- tural changes during ESI. Thus, there is a critical need to understand how biomolecular structures are modified during and after the ESI process to determine (1) how researchers can deduce solvated structures from these techniques and (2) how analytical methods can be improved. To address this challenge, the Gallagher lab uses a combination of molecular dynamics (MD) simulations and MS methods to develop a fundamental, molecular perspective of ESI. The Gallagher lab performs MD simulations of ESI to observe carbohydrate ionization using metal ions as charge carriers. However, to more accurately model protein ionization, the Gallagher lab is developing methods to simulate protonation during ESI. These methods will be further developed to examine analyte ionization by deprotonation in negative-ion mode, and then applied to achieve a molecular perspective on the ionization of glycans and proteins. In paral- lel to the MD simulations, the Gallagher lab will perform MS analysis of metal-adducted glycans. Glycans readily ionize by coordinating to metal ions, with different metal-adducts enabling isomeric differentiation in both tandem MS and ion mobility-MS. However, glycans are often analyzed as sodium adducts because so- dium is a ubiquitous contaminant. The Gallagher lab is performing systematic studies examining the relation- ship between metal-ion properties and glycan characterization by MS. Finally, the Gallagher lab is applying native MS to characterize protein complexes. In native MS, noncovalent interactions are maintained in the gas phase; yet past work has suggested that proteins analyzed as positive versus negative ions have differ- ences in gas-phase stability. The Gallagher lab will examine how charging in ESI to form either positive or neg- ative ions is related to gas-phase protein structure, stability, and ligand-binding interactions. The overarching goal of the Gallagher lab is to develop and apply novel methodologies to characterize gly- cans, proteins, and their binding interactions. The research described in this proposal is significant because it will provide a fundamental perspective on ESI-MS and the molecular insights from this research will enable the rational development of future ESI-MS methods for characterizing these molecules. Ultimately, this work will facilitate the analysis of glycans, glycoconjugates, and protein complexes in cell biology and disease states, enabling these methods to be applied to address important biological hypotheses.

项目概要/摘要 聚糖是碳水化合物的异质聚合物，与蛋白质相互作用，引发多种生物活性 钙过程，包括分子识别、细胞信号传导和宿主-微生物相互作用。质谱- 旋转测量 (MS) 方法已成为表征聚糖结构及其相互作用的有力工具。 与蛋白质的作用。电喷雾电离 (ESI) 是一种常见的电离方法，用于转移这些生物 分子从溶液到气相进行 MS 分析。然而，聚糖和蛋白质都表现出结构 ESI 期间的自然变化。因此，迫切需要了解生物分子结构是如何修饰的 在 ESI 过程期间和之后确定 (1) 研究人员如何从中推断出溶剂化结构 技术以及（2）如何改进分析方法。 为了应对这一挑战，Gallagher 实验室结合使用了分子动力学 (MD) 模拟 和 MS 方法来开发 ESI 的基本分子视角。 Gallagher 实验室执行 MD ESI 模拟以观察使用金属离子作为电荷载体的碳水化合物电离。然而，为了更多 为了准确地模拟蛋白质电离，加拉格尔实验室正在开发模拟质子化过程的方法 ESI。这些方法将进一步发展，以通过负离子中的去质子化来检查分析物的电离 模式，然后应用于实现聚糖和蛋白质电离的分子视角。并列中—— 与 MD 模拟相比，Gallagher 实验室将对金属加合聚糖进行 MS 分析。聚糖 通过与金属离子配位而容易电离，不同的金属加合物可以实现异构体分化 串联 MS 和离子淌度 MS。然而，聚糖通常被分析为钠加合物，因为 - 镉是一种普遍存在的污染物。加拉格尔实验室正在进行系统研究，检验以下关系： 通过 MS 在金属离子特性和聚糖表征之间进行转换。最后，加拉格尔实验室正在申请 天然 MS 来表征蛋白质复合物。在天然 MS 中，非共价相互作用维持在 气相；然而过去的研究表明，以正离子和负离子分析的蛋白质具有不同的特性。 气相稳定性的影响。加拉格尔实验室将研究如何在 ESI 中充电以形成正电或负电 活性离子与气相蛋白质结构、稳定性和配体结合相互作用有关。 加拉格尔实验室的首要目标是开发和应用新的方法来表征gly- 罐头、蛋白质及其结合相互作用。本提案中描述的研究意义重大，因为它 将为 ESI-MS 提供基本视角，并且本研究的分子见解将使 合理开发未来 ESI-MS 方法来表征这些分子。最终，这项工作将 促进细胞生物学和疾病状态中聚糖、糖复合物和蛋白质复合物的分析， 使这些方法能够应用于解决重要的生物学假设。