Salt-induced fibrillogenesis of fibrinogen (SAL-FIB): In vitro experiments and simulations

盐诱导纤维蛋白原原纤维形成 (SAL-FIB)：体外实验和模拟

• 批准号: 462381005
• 负责人: Professorin Dr. Dorothea Brüggemann
• 金额: --
• 依托单位: Bremen Center for Computational Materials Science (BCCMS)
• 依托单位国家: 德国
• 项目类别: Research Grants
• 资助国家: 德国
• 项目状态: 未结题
• 来源: https://gepris.dfg.de/gepris/projekt/462381005?language=en
• 关键词: Salt; induced; fibrillogenesis; fibrinogen; SAL

Fibrous fibrinogen scaffolds are particularly attractive for tissue engineering applications since they closely mimic the architecture and biochemical composition of native blood clots. Different techniques are known to induce fibrillogenesis of fibrinogen under in vitro conditions including electrospinning, surface- and buffer-driven fiber formation as well as salt-induced self-assembly. However, to date it is not yet understood whether specific surface interactions or buffer conditions contribute to the in vitro fibrillogenesis of fibrinogen. Moreover, current studies on the molecular level of fibrinogen lack a clear understanding of the underlying atomistic processes. Therefore, our project will focus on the following question: Which mechanism drives fibrillogenesis of fibrinogen under in vitro conditions? To answer this question our project will use a multi-scale combination of closely connected simulative and experimental methods. For the first time, we will establish a complete molecular model of fibrinogen, that also includes posttranslational modifications. Based on experimental dynamic light scattering studies, where we will analyze the aggregation of fibrinogen in different buffer systems, this fibrinogen model will be used in molecular dynamics (MD) studies to analyze local ion distributions around the molecule. Subsequently, we will use our MD results to study whether salt ions are incorporated into fibrinogen molecules or into fibrinogen fibers upon drying. The associated experimental studies will involve turbidimetric measurements and analysis of the morphology and elemental composition of self-assembled fibrinogen nanofibers. To understand whether fiber assembly is accompanied or driven by changes in the fibrinogen conformation, we will screen self-assembled fibers for possible amyloid transitions and perform spectroscopic analyses. The resulting circular dichroism (CD) spectra will be compared to reference single molecule CD spectra with induced structural changes obtained by steered MD simulations.A major challenge in this project will be to discuss and interpret the single molecule information from MD simulations with respect to experimental results obtained from an ensemble of fibrinogen structures/conformations. In this context, the hydrodynamic radius and CD spectra will be two observables that will directly link our simulative and experimental studies. Only with this combined approach we will be able to propose a detailed mechanism for the in vitro fibrillogenesis of fibrinogen with a clear dependence on environmental parameters. The proposed project will therefore provide fundamental insights into in vitro fibrillogenesis of fibrinogen, which are necessary to develop a new class of fibrinogen nanofibers with defined structure-function-relationships.

纤维纤维蛋白原支架对于组织工程应用特别有吸引力，因为它们紧密地模仿天然血凝块的结构和生化组成。已知不同的技术在体外条件下诱导纤维蛋白原的原纤维形成，包括静电纺丝、表面和缓冲液驱动的纤维形成以及盐诱导的自组装。然而，到目前为止，它还不清楚是否特定的表面相互作用或缓冲条件有助于纤维蛋白原的体外原纤维形成。此外，目前对纤维蛋白原分子水平的研究缺乏对潜在原子过程的清晰理解。因此，我们的项目将集中在以下问题：在体外条件下，什么机制驱动纤维蛋白原的原纤维形成？为了回答这个问题，我们的项目将使用紧密相连的模拟和实验方法的多尺度组合。我们将首次建立一个完整的纤维蛋白原分子模型，其中也包括翻译后修饰。基于实验动态光散射研究，我们将分析纤维蛋白原在不同缓冲系统中的聚集，该纤维蛋白原模型将用于分子动力学（MD）研究，以分析分子周围的局部离子分布。随后，我们将使用我们的MD结果来研究盐离子是否掺入纤维蛋白原分子或干燥后的纤维蛋白原纤维。相关的实验研究将涉及浊度测量和自组装纤维蛋白原纳米纤维的形态和元素组成的分析。为了了解纤维蛋白原构象的变化是否伴随或驱动纤维组装，我们将筛选自组装纤维中可能的淀粉样蛋白转变并进行光谱分析。将所得的圆二色性光谱与由分子动力学模拟得到的具有诱导结构变化的参考单分子圆二色性光谱进行比较，并结合纤维蛋白原结构/构象系综的实验结果讨论和解释分子动力学模拟得到的单分子信息。在这种情况下，流体动力学半径和CD光谱将是两个观测值，将直接连接我们的模拟和实验研究。只有通过这种结合的方法，我们将能够提出一个详细的机制，在体外纤维蛋白原与环境参数的明确依赖。因此，拟议的项目将提供基本的见解，在体外纤维蛋白原，这是必要的开发一类新的纤维蛋白原纳米纤维与定义的结构-功能-关系。