An integrated mass spectrometry approach to study heparin structure-bioactivity

研究肝素结构-生物活性的综合质谱方法

• 批准号: 9252476
• 负责人: IGOR A KALTASHOV
• 金额: $ 31.19万
• 依托单位: UNIVERSITY OF MASSACHUSETTS AMHERST
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-04-01 至 2020-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9252476
• 关键词: Address; Adopted; Affinity; Alzheimer' s Disease; Anticoagulants; Antithrombin III; Area; Binding; Binding Sites; Biopolymers; Blood coagulation; Cell Adhesion; Cell Differentiation process; Cell Proliferation; Charge; Chemistry; Clinic; Clinical; Complex; Crystallization; Data; Detection; Deuterium; Digestion; Disease; Dissociation; Drug Delivery Systems; Electrons; Electrostatics; Embryonic Development; Exhibits; Family; Fibroblast Growth Factor; Gases; Glycosaminoglycans; Goals; Heparin; Heparin Lyase; Heparinoids; Heterogeneity; Hydrogen; Inflammation; Ion-Exchange Chromatography Procedure; Ions; Kinetics; Knowledge; Length; Malignant Neoplasms; Mass Spectrum Analysis; Measures; Medical; Medicine; Methodology; Modality; Modeling; Molecular; Molecular Conformation; Molecular Models; Monte Carlo Method; Multiple Partners; Neoplasm Metastasis; Neurodegenerative Disorders; Parkinson Disease; Pathologic; Pharmacology; Phase; Physiological; Physiological Processes; Polymers; Polysaccharides; Process; Property; Proteins; Regenerative Medicine; Research; Resolution; Role; Source; Spectrometry, Mass, Electrospray Ionization; Structure; Techniques; Therapeutic; Therapeutic Agents; Therapeutic Effect; Tissue Engineering; Vertebral column; Work; Wound Healing; angiogenesis; base; cationic antimicrobial protein CAP 37; clinical application; design; holistic approach; immunogenic; immunoregulation; improved; ion mobility; member; molecular modeling; novel; oncology; protein complex; public health relevance; scaffold; simulation; stoichiometry; structural biology; therapeutic target; tissue regeneration; tool

 DESCRIPTION (provided by applicant): Heparin-related glycosaminoglycans (GAGs) hold enormous promise in the field of regenerative medicine, and also are a largely unexploited source of therapeutic agents to address a range of other medically important conditions including cancer and neurodegenerative diseases. Nevertheless, their practical utilization in the clinic is still confined to anticoagulants, while progress in other areas remains relatively modest The great challenge presented by these biopolymers to medicine and, more broadly, to structural biology is due in large part to their enormous structural heterogeneity, which is mirrored by their multiple roles in modulating angiogenesis, cell adhesion, embryogenesis, inflammation, metastasis and wound healing. Despite extensive research efforts, clear understanding of how various structural features modulate function of these biopolymers remains wanting, arguably because the inquiry was designed using the framework of the lock-and-key model, a paradigm that had been tremendously successful in structural biology applied to proteins. However, this approach fails to recognize that the dramatically higher level of structural diversity exhibited by GAGs has functional importance. The proposed research seeks to shift the prevailing paradigm in the field of GAG/protein interactions by moving away from the notion of well-defined sterically complementary binding sites and emphasizing a greater role of polyvalent electrostatics. In addition to revealing the determinants of GAG/protein interactions and understanding how they fine-tune the binding process, we will obtain a high-resolution and dynamic description of the mechanism of heparin interaction with several therapeutically relevant proteins. This will be done without either limiting the structural space to a few precisel defined GAG molecules or reducing the conformational space to a few static snapshots available from crystal structures. Towards this goal, we will adopt a "holistic" approach to protein/GAG interactions that embraces the structural diversity of heparin-like GAGs, as opposed to the commonly accepted approach which limits the scope of inquiry to a few molecules accessible through synthesis. We will use a combination of bottom-up and top-down approaches to study protein/GAG interactions, the former focusing on relatively well-defined subsets of short heparin oligomers obtained by affinity separations, and the latter aimed at intact heparin. This will reveal the structural properties governing the interactions of GAGs with several therapeutically relevant proteins. Novel applications of gas-phase ion chemistry (electron capture and collisional activation) will be developed for characterization of large heterogeneous GAG/protein complexes. Finally, atomic-level Monte Carlo simulations in parallel with H/D exchange studies will lead to a high-resolution and dynamic depiction of the interaction process. This new view of GAG/protein interactions will allow the therapeutic potential of heparin-like GAGs to be exploited more efficiently in areas as diverse as regenerative medicine, inflammation, neurodegenerative disorders and oncology.

 描述（由申请人提供）：肝素相关糖胺聚糖（GAG）在再生医学领域具有巨大的前景，也是治疗一系列其他医学重要疾病（包括癌症和神经退行性疾病）的治疗剂的主要未开发来源。然而，它们在临床上的实际应用仍然局限于抗凝剂，而在其他领域的进展仍然相对温和。这些生物聚合物对医学以及更广泛地说对结构生物学提出的巨大挑战在很大程度上是由于它们巨大的结构异质性，这反映在它们在调节血管生成、细胞粘附、胚胎发生、炎症、转移和伤口愈合中的多重作用。尽管进行了广泛的研究工作，但仍然需要清楚地了解各种结构特征如何调节这些生物聚合物的功能，可以说是因为调查是使用锁和钥匙模型的框架设计的，这是一种在结构生物学中应用于蛋白质的范式。然而，这种方法未能认识到，糖胺聚糖所表现出的显著更高水平的结构多样性具有功能重要性。拟议的研究旨在通过远离定义明确的空间互补结合位点的概念并强调多价静电的更大作用来改变GAG/蛋白质相互作用领域的流行范式。除了揭示GAG/蛋白质相互作用的决定因素并了解它们如何微调结合过程外，我们还将获得肝素与几种治疗相关蛋白质相互作用机制的高分辨率和动态描述。这将在不将结构空间限制为几个精确定义的GAG分子或将构象空间减少到几个可从晶体结构获得的静态快照的情况下完成。为了实现这一目标，我们将采用一种“整体”的方法来蛋白质/GAG相互作用，包括肝素样GAG的结构多样性，而不是普遍接受的方法，将调查范围限制在通过合成可获得的几个分子。我们将结合使用自下而上和自上而下的方法来研究蛋白质/GAG相互作用，前者侧重于通过亲和分离获得的相对明确的短肝素寡聚体子集，后者旨在研究完整的肝素寡聚体。 肝素这将揭示控制GAG与几种治疗相关蛋白质相互作用的结构特性。气相离子化学（电子捕获和碰撞活化）的新应用将开发用于表征大的异质性GAG/蛋白质复合物。最后，原子级蒙特卡罗模拟与H/D交换研究并行将导致相互作用过程的高分辨率和动态描述。GAG/蛋白质相互作用的这种新观点将使肝素样GAG的治疗潜力在再生医学、炎症、神经退行性疾病和肿瘤学等领域得到更有效的利用。