Force-induced Conformational Transitions in Single Polysaccharide Molecules by AFM

由基于AFM吡喃糖环的分子在单个多糖分子中的力诱导的构象转变对于生物系统（例如葡萄糖）具有特别重要的意义。许多重要的生物多糖都是由吡喃糖环组成的，它们在各种细胞结构如植物的细胞壁或动物组织的细胞外基质（ECM）中处于张应力下。机械应力被认为调节这些复杂弹性系统的组装和生理特性。多糖通过其组分的机械重排来应对这种压力，但其潜在机制尚不清楚。最简单的观点认为多糖是熵弹簧，吡喃糖环结构通常被描绘成无弹性的，并被锁定在一个稳定的构象中。单分子原子力显微镜技术的发展允许严格审查这些观点。原子力显微镜仪器可以机械拉伸单分子，并具有极好的长度和力分辨率。用原子力显微镜对多糖进行拉伸实验，发现多糖并不表现为简单的熵弹簧，而是表现出屈服现象。这些弹性偏差的起源被精确地指向吡喃糖环，该吡喃糖环被发现在拉伸时经历构象转变，如椅船或椅反转。这些强制跃迁以逐步的方式改变了糖苷氧原子的分离，从而影响了多糖链的轮廓长度及其弹性。轴向糖苷键被发现通过充当原子杠杆来驱动这些转变。这个建议的长期目标是在原子水平上理解多糖中这些力诱导的构象转变的机制。在第二个资助期内，单分子原子力显微镜技术将与计算化学的工具结合起来，详细研究力诱导的吡喃糖环构象转变。环的机械性能和它的机械构象转变将通过从不同方向和不同的连接点拉动环来探测。 这些研究将确定每种类型的单体对混合链的复合弹性的贡献，并将在解释许多天然多糖的分子弹性方面具有价值。该建议的实验和理论研究结果将被整合，以开发一种基于AFM的方法，用于从其独特的力延伸光谱中识别溶液中的单个多糖分子。这样的方法将是一个重要的补充，以分析工具库碳水化合物的研究。

Force-induced conformational transitions in single polysaccharide molecules by AFMPyranose ring-based molecules are of extraordinary importance to biological systems (e.g.glucose). Many biologically important polysaccharides are composed of pyranose rings and theyare placed under tensile stress in a wide variety of cellular structures such as the cell wall ofplants or the extracellular matrix (ECM) in animal tissues. Mechanical stress is thought toregulate assembly and physiological properties of these complex elastic systems. Thepolysaccharides respond to that stress by mechanical rearrangements of their components butthe underlying mechanism is not well understood. The simplest view assumes thatpolysaccharides are entropic springs and the pyranose ring structure is typically portrayed asinelastic and locked into a stable conformation. The development of single molecule AFMtechniques allowed for critically examining these views. AFM instruments can stretchmechanically single molecules and have superb length and force resolution. Stretching ofpolysaccharides by AFM revealed that they do not behave as simple entropic springs but thatthey display yielding phenomena. The origin of these elastic deviations was pinpointed to thepyranose ring that was found to undergo, upon stretching, conformational transitions such aschair-boat or chair inversion. These forced transitions change, in a step-wise fashion, theseparation of the glycosidic oxygen atoms, and therefore affect the contour length of thepolysaccharide chain and its elasticity. Axial glycosidic bonds were found to drive thosetransitions by acting as atomic levers. The long-term objective of this proposal is to understand,at the atomic level, the mechanism of these force-induced conformational transitions inpolysaccharides. During the second grant period single molecule AFM techniques will becombined with the tools of computational chemistry to examine in detail force-inducedconformational transitions in the pyranose ring. The mechanical properties of the ring and itsmechanical conformational transitions will be probed by pulling on the ring from variousdirections and by different attachment points. These studies will determine the contribution of each type of the monomer to thecomplex elasticity of a mixed chain and will be valuable in interpreting the molecular elasticity ofmany native polysaccharides. The experimental and theoretical findings of this proposal will beintegrated to develop an AFM-based methodology for identifying individual polysaccharidemolecules in solution from their unique force-extension spectra. Such a methodology will be animportant addition to the arsenal of analytical tools for carbohydrate research.