超声和金属离子对胰高血糖素聚集的影响

发布时间：2018-03-17 14:01

  本文选题：胰高血糖素　切入点：超声　出处：《宁波大学》2012年硕士论文　论文类型：学位论文

【摘要】：蛋白的淀粉样纤维化聚集与多种疾病密切相关，如老年痴呆症等。研究发现，蛋白纤维的聚集不仅和氨基酸的序列有关，而且还与外界环境因素如金属离子、pH、超声、温度等相关。在本文中，我们的工作主要包含以下两个方面：（1）研究胰高血糖素在超声波影响下的聚集过程。在此工作中，我们利用原子力显微镜(AFM)观察蛋白聚集的形貌，并结合硫黄素(thioflavin T，，ThT)荧光检测其形成淀粉样纤维的动力学曲线，发现在超声波的影响下，胰高血糖素可在较短时间内开始形成淀粉样纤维，大大缩短了蛋白形成纤维的“酝酿”时间。而且在不同超声功率和不同超声频率下形成的纤维形貌有较大差异。并用显微傅立叶变换红外光谱（Micro-FTIR）对超声聚集而成的蛋白纤维的二级结构进行表征。结果发现此过程中，蛋白的二级结构发生了转变(α螺旋到β折叠)。（2）研究金属离子对胰高血糖素淀粉样蛋白聚集造成的影响。我们主要是利用原子力显微镜和显微傅立叶变换红外光谱对加入Fe(Ⅲ)、Cu(Ⅱ)、Fe(Ⅱ)后的胰高血糖素进行表征，结果发现加入Fe(Ⅲ)后，胰高血糖素蛋白聚集形成淀粉样纤维的过程进行得非常缓慢，而且，经过孵化的淀粉样蛋白的表面形貌以及它的二级结构都大大地改变了；但是对于加入Cu(Ⅱ)的样品，胰高血糖素进行纤维化的程度却变得较快，所得到的表面形貌也显然不同，不过淀粉样纤维的二级结构却没怎么有大的改变；对于掺有Fe(Ⅱ)的蛋白，纤维化过程在速度上没怎么变化，不过二级结构却变了。此外，我们还利用了圆二色谱（CD）和ThT荧光进一步对加入Fe(Ⅲ)、Cu(Ⅱ)后胰高血糖素的聚集过程进行研究，关于其二级结构的变化与FTIR得出的结论相同，荧光动力学曲线显示出加入Fe(Ⅲ)后，蛋白聚集的进程被抑制，而加有Cu(Ⅱ)的胰高血糖素的纤维化速度却在一定程度上提高了。
[Abstract]:The accumulation of amyloid fibrosis is closely related to many diseases, such as Alzheimer's disease. Studies have found that the aggregation of protein fibers is not only related to the sequence of amino acids, but also to environmental factors such as metal ion pH, ultrasound, etc. In this paper, we mainly study the aggregation process of glucagon under the influence of ultrasonic wave. In this work, we use AFM (atomic force microscope) to observe the morphology of protein aggregation. The kinetic curves of the formation of amyloid fibers were determined by fluorescence analysis of thioflavin Th Th. It was found that glucagon could form amyloid fibers in a short time under the influence of ultrasound. The "brewing" time of protein forming fiber was shortened greatly, and the morphology of the fiber formed under different ultrasonic power and frequency was different. Micro-FTIR was used to aggregate the ultrasonic. The secondary structure of the protein fibers was characterized. The secondary structure of the protein was transformed (伪 -helix to 尾 -fold) to study the effect of metal ions on the aggregation of glucagon amyloid protein. We mainly use atomic force microscope (AFM) and micro-Fourier transform infrared spectroscopy (FT-IR) to study the effect of metal ions on the aggregation of glucagon amyloid protein. The glucagon was characterized by adding Fe (鈪

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