Federal Research and Clinical Center of Physical-Chemical Medicine, Malaya Pirogovskaya, 1a, Moscow 119435 Russian Federation.
Federal Research and Clinical Center of Physical-Chemical Medicine, Malaya Pirogovskaya, 1a, Moscow 119435 Russian Federation; University of Pennsylvania School of Medicine, Department of Cell and Developmental Biology, 421 Curie Boulevard, Philadelphia, PA 19104, USA.
Colloids Surf B Biointerfaces. 2018 Jul 1;167:370-376. doi: 10.1016/j.colsurfb.2018.04.037. Epub 2018 Apr 19.
Fibrinogen denaturation is an important phenomenon in biology and medicine. It has been previously investigated with bulk methods and characterized by parameters, which refer to big protein ensembles. Here we provide a new insight into fibrinogen denaturation with a high-resolution single-molecule atomic force microscopy (AFM). The ultrastructure of individual fibrinogen molecules was studied after heating or extended contact with the highly oriented pyrolytic graphite surface (HOPG) modified with oligoglycine-hydrocarbon graphite modifier (GM). Fibrinogen heating to 65 °C and 90 °C resulted in the formation of various shapes containing fibrillar and globular structures, which were attributed to the monomers and small aggregates of fibrinogen. Fibrinogen unfolded by the extended (10 min) incubation on GM-HOPG surface in water revealed a different morphology. It contained fibrillar structures only, and their organization reflected the initial native structure of fibrinogen: typically, six polypeptide chains connected by multiple disulfide bonds were seen. A combination of two morphologies - globular aggregates with dense fibrillar networks - has been revealed for thermally denatured protein adsorbed on a GM-HOPG surface with extended (10 min) rinsing with water. The obtained results provide better understanding of fibrinogen unfolding induced by different factors and are important for improvement of biomedical applications, such as fibrinogen-based protein matrixes and carbon-based biomaterials.
纤维蛋白原变性是生物学和医学中的一个重要现象。它以前曾用体相方法进行过研究，并通过参数来进行表征，这些参数涉及到大的蛋白质集合体。在这里，我们通过高分辨率单分子原子力显微镜（AFM）为纤维蛋白原变性提供了新的见解。在加热或与高度取向的热解石墨表面（HOPG）长时间接触后，研究了经过寡甘氨酸-碳氢化合物石墨修饰剂（GM）修饰的单个纤维蛋白原分子的超微结构。将纤维蛋白原加热至 65°C 和 90°C 会导致形成各种形状，其中包含纤维状和球状结构，这些结构归因于纤维蛋白原的单体和小聚集体。在水中与 GM-HOPG 表面长时间（10 分钟）孵育导致纤维蛋白原展开，呈现出不同的形态。它只含有纤维状结构，其组织反映了纤维蛋白原的初始天然结构：通常可以看到由多个二硫键连接的六个多肽链。在 GM-HOPG 表面上吸附的热变性蛋白质在经过 10 分钟的水冲洗后，其两种形态（密集的纤维状网络的球形聚集体）的组合被揭示出来。这些结果提供了对不同因素诱导的纤维蛋白原展开的更好理解，对于改进生物医学应用（例如基于纤维蛋白原的蛋白质基质和基于碳的生物材料）非常重要。
