利用离子淌度质谱解析配体对蛋白质结构的影响
发布时间：2019-04-03 19:30
【摘要】：离子淌度质谱(ion mobility mass spectrum,IM-MS)即将离子淌度分离技术与质谱联用的一项新型分析方法,已成为一种强有力的结构生物学分析工具,能同时测定分析物的结构、拓扑学结构、动态过程以及组成等。本论文以膜联蛋白A5及其同质二聚体diannexin和维生素B12结合蛋白F为材料,探究不同配体如钙离子和维生素B12对蛋白质结构的影响。由电喷雾电离产生的蛋白质离子在离子淌度质谱仪的漂移管中与气体发生碰撞而导致蛋白质被活化,促使其构象失去天然状态发生去折叠等变化,且离子在漂移管中会根据他们的质量、电荷、大小被逐渐分离。在本论文中我们采用Waters Synapt G2-Si HDMS质谱仪采集离子的漂移时间,并经过许多已知理论碰撞横截面积的蛋白质标准品做校正后得到离子天然构象与去折叠状态的碰撞横截面积。实验发现:未受碰撞活化的离子,其测得的碰撞横截面积与预期的处于天然状态的蛋白质的碰撞横截面积非常一致。而当碰撞能量增加,离子被活化导致构象发生变化,且构象去折叠的程度与蛋白质复合物分子内或分子间的非共价相互作用的稳定性有关。以上结果表明蛋白质与配体结合后能使其在气相中的结构稳定性大大增强。
[Abstract]:Ion mobility mass spectrometry (ion mobility mass spectrum,IM-MS), which combines ion mobility separation with mass spectrometry, has become a powerful tool for structural biological analysis, which can simultaneously determine the structure of analytes. Topological structure, dynamic process and composition, etc. In this paper, the effects of different ligands, such as calcium ion and vitamin B12, on protein structure were investigated by using membrane protein A5 and its homodimer diannexin and vitamin B12 binding protein F as materials. The protein ion produced by electrospray ionization collides with the gas in the drift tube of the ion mobility mass spectrometer, which causes the protein to be activated, causing its conformation to lose its natural state and unfold and so on. And ions in the drift tube will be gradually separated according to their mass, charge, and size. In this paper, we use Waters Synapt G2-Si HDMS mass spectrometer to collect the drift time of ion, and after correction of many protein standards with known theoretical collision cross-sectional area, we obtain the collision cross-sectional area of ion natural conformation and unfolding state. It is found that the collision cross-sectional area measured by the ion which is not activated by collision is in good agreement with the expected collision cross-sectional area of the protein in the natural state. When the collision energy increases, the ion activation causes the conformation change, and the degree of conformation unfolding is related to the stability of intramolecular or intermolecular noncovalent interactions of protein complexes. The above results indicate that the structure stability of the protein in gas phase can be greatly enhanced by binding the protein with ligand.
【学位授予单位】：南京理工大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：O657.63;TQ931
[Abstract]:Ion mobility mass spectrometry (ion mobility mass spectrum,IM-MS), which combines ion mobility separation with mass spectrometry, has become a powerful tool for structural biological analysis, which can simultaneously determine the structure of analytes. Topological structure, dynamic process and composition, etc. In this paper, the effects of different ligands, such as calcium ion and vitamin B12, on protein structure were investigated by using membrane protein A5 and its homodimer diannexin and vitamin B12 binding protein F as materials. The protein ion produced by electrospray ionization collides with the gas in the drift tube of the ion mobility mass spectrometer, which causes the protein to be activated, causing its conformation to lose its natural state and unfold and so on. And ions in the drift tube will be gradually separated according to their mass, charge, and size. In this paper, we use Waters Synapt G2-Si HDMS mass spectrometer to collect the drift time of ion, and after correction of many protein standards with known theoretical collision cross-sectional area, we obtain the collision cross-sectional area of ion natural conformation and unfolding state. It is found that the collision cross-sectional area measured by the ion which is not activated by collision is in good agreement with the expected collision cross-sectional area of the protein in the natural state. When the collision energy increases, the ion activation causes the conformation change, and the degree of conformation unfolding is related to the stability of intramolecular or intermolecular noncovalent interactions of protein complexes. The above results indicate that the structure stability of the protein in gas phase can be greatly enhanced by binding the protein with ligand.
【学位授予单位】：南京理工大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：O657.63;TQ931
【参考文献】
相关期刊论文 前2条
1 ;Identification of Bovine Casein Phosphorylation Using Titanium Dioxide Enrichment in Combination with Nano Electrospray Ionization Tandem Mass Spectrometry[J];Journal of Integrative Agriculture;2012年03期
2 张立勇,赵晓航,吴e，
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