采用分子动力学模拟和简化蛋白质模型研究人微球蛋白83-89片段低聚体的自组装过程

发布时间：2018-01-16 01:19

本文关键词：采用分子动力学模拟和简化蛋白质模型研究人微球蛋白83-89片段低聚体的自组装过程　出处：《复旦大学》2008年硕士论文　论文类型：学位论文

【摘要】： 神经细胞中的特定蛋白质呈淀粉样聚集会导致包括老年痴呆症,帕金森氏症以及疯牛病在内的超过20种神经性退化疾病。本文中所探讨的人微球蛋白也会产生相似的聚集过程而导致血液透析病。在过去20年中,尽管人们已经做了大量的实验和理论工作,但是无论对于完整致病蛋白质还是其中的某些片段,其聚集早期阶段的详细过程和机理并没有被研究透彻.对聚集早期阶段的研究正引起研究人员的浓厚兴趣。因为不断有证据表明聚集过程中水溶性的低聚体比完整的纤维状态具有更强的细胞毒性,并且这些由不同致病蛋白或蛋白片段形成的低聚体可以被相同的抗体识别。研究蛋白质聚集的初始阶段不仅可以从分子角度了解低聚体形成的过程,而且可以揭示不同的氨基酸序列对于聚集动力学和低聚体结构的影响。虽然科学家已经发现很多种蛋白质可以聚集形成纤维化结构,但是在分子层面上,由于纤维结构的不可溶性和实验手段的制约,人们对蛋白质纤维低聚体的结构和聚集过程的了解仍然有限。由于链状分子折叠的过程复杂,通常熵的变化会对结构产生较大影响,同时低聚体聚集过程发生在纳米的空间尺度和纳秒至微秒的时间尺度,使大多数实验检测手段受到限制。因此计算机模拟可以弥补实验方面的不足,成为研究多肽聚集问题的有效方法。蛋白质聚集形成的淀粉样纤维有一些共同的特征,包括在显微镜下具有相似的形貌、可以与染色剂刚果红结合、以及具备Cross-β特征结构的X射线衍射图案。蛋白质聚集研究中普遍关注的问题包括了Cross-β的具体结构、形成纤维的基本结构模块(building blocks)、氨基酸序列对结构模块的影响以及低聚体聚集的成核过程。本文中所介绍的工作,是采用基于简化蛋白质模型OPEP(Optimized Potential for Efficient peptide structure Prediction)力场的分子动力学模拟,研究人的微球蛋白83-89片段的低聚体聚集过程,描述了四聚体和七聚体的自由能面以及16聚体聚集的初始阶段。特别是对结构模块的具体形态,低聚体的成核过程等问题进行了详细的讨论和研究。本文阐述的主要成果分为以下几个部分: 1.在第三章中,我们针对可以产生淀粉样聚集的人微球蛋白83-89片段的四聚体进行初步研究。由于此前对于该片段的计算机研究只达到二聚体的大小,我们同时采用分子动力学(Molecular Dynamics,MD)和蒙特卡罗(Monte Carlo,MC)方法研究四聚体在纤维聚集初期可能出现的聚集形态,以及直接的热力学性质和动力学转化过程。根据成核理论,蛋白质聚集过程中成核尺寸可能随片段序列的不同而存在差异。我们发现,对于人微球蛋白83-89片段低聚体,模拟得到的可能形成纤维结构的有序构形几率都比较低,由此可以推断该片段的成核尺寸要多于四条链。与此同时,模拟中还发现了卷曲的单层β折叠和环状结构。在后续研究中我们发现,这些结构作为形成更大更有序构形的过渡状态而存在。 2.在第四章中,为检验在前期四聚体模拟中得到的具有纤维基本模块特征的两种结构,即平展单层β折叠和双层β折叠是否具有进一步形成结构模块的可能性,我们模拟了83-89片段的七聚体体系。研究发现,随着体系增大,即链的数目增加,疏水相互作用开始对结构的形成以及稳定产生至关重要的作用。单层的β折叠不复存在,相应的平行双层β折叠几率明显上升,并且在层与层之间形成了稳定的疏水基团。从我们的模拟结果推断,平行双层β斤叠结构是人微球蛋白83-89片段纤维模块可能的聚集构形。在模拟过程中,我们还发现了β桶状结构,即由β折叠卷曲包围形成的通道结构,在桶的中心存在紧密的疏水核,由非极性氨基酸侧链原子组成。这种聚集过程出现的桶状可以通过两种路径形成。一种是由卷曲单层β折叠通过结合游离单链加长围成;另一种是直接通过垂直的双层β折叠转化而成。深入研究β桶状结构可以发现,它可以与垂直双层β折叠结构相互转化,进而转化成平行双层结构。所以我们认为,β桶状结构是低聚体聚集过程中的中间状态。 3.第五章的主要内容是在计算资源和模拟时间允许的条件下,结合所采用的MD-OPEP方法具有高效率的特点,进行83-89片段16聚体的分子动力学模拟。在已有的6个模拟中,体系并没有直接形成有序结构模块的形态。出现的多种多样的球形空间结构说明,在疏水相互作用强于氢键相互作用时,体系需要更长的聚集时间形成稳定而有序的氢键,从而形成稳定的纤维结构模块。 4.最后,在第六章中,总结了前文中针对人微球蛋白83-89片段不同体系所进行的研究。通过类比这三种不同大小的体系,我们发现了83-89片段低聚体的一系列有序构形、形成并维持这些构形的主要相互作用类型、纤维前体的结构模块、特殊的转变中间态以及聚集成核的过程。
[Abstract]:Nerve cell specific proteins in amyloid aggregation causes including Alzheimer's disease, Parkinson's disease, more than 20 kinds of mad cow disease and neural degenerative disease. This article discussed the human microglobulin will produce similar aggregation process caused by hemodialysis patients. In the past 20 years, although people have to do a large amount of experimental and theoretical work, but for some fragments of pathogenic protein or the complete, detailed process and mechanism of the accumulation of early stage has not been studied thoroughly. It resulted in the early stages of the aggregation of interested researchers. Because there is growing evidence that the cytotoxicity of water soluble oligomers of the aggregation process the fiber is stronger than the complete state, and these oligomers by different pathogenic proteins or protein fragments can be formed by the same antibody. Study on protein The initial stage of mass aggregation can not only understand the formation process of oligomers from molecular perspective, but also reveal the effects of different amino acid sequences on aggregation kinetics and oligomer structure.
Although scientists have found many kinds of proteins can be aggregated to form fibrous structure, but at the molecular level, due to the restriction of fiber structure is not soluble and experimental means, people on the structure of oligomeric protein fiber and aggregation process understanding is still limited. Because of the complex molecule folding, changes of entropy will have great influence on at the same time, oligomeric structure, aggregation occurs in the nano space scale and nanosecond to microsecond time scale, make the most of the experimental detection means is limited. So the computer simulation can make up for lack of experimental aspects, is an efficient method to study the problem of aggregation of peptides.
The amyloid protein aggregation formation have some common features, including with similar morphology under the microscope, and dye Congo red binding, as well as X ray diffraction pattern characteristic structure of Cross- beta protein aggregation. Attention problems in the study include the specific structure of Cross- beta, form the basic structure of the fiber module (building blocks). The effect of amino acid sequence on the structure of module and the oligomeric aggregation nucleation process. This paper introduced in the work, is a simplified protein model based on OPEP (Optimized Potential for Efficient peptide structure Prediction) molecular dynamics simulation of stress field, the process of oligomeric protein microspheres study of human 83-89 fragment accumulation describes the four dimers and seven dimers of the free energy surface and 16 dimer aggregation of the initial stage. Especially the specific form of the structure module, low The nucleation process of polymer is discussed and studied in detail. The main achievements of this paper are divided into the following parts:
1. in the third chapter, we make a preliminary study on four dimers can produce amyloid aggregation of human microglobulin 83-89 fragment. The fragment due to previous computer research only reached the size of two dimers, we use molecular dynamics (Molecular, Dynamics, MD) and Monte Carlo (Monte Carlo MC) method form four dimers in the fiber aggregation aggregation may occur early, and direct the thermodynamic and kinetic conversion process. According to nucleation theory, the protein aggregation process of nucleation size may vary with the sequence differences. We found that the human microglobulin fragment of 83-89 oligomers, ordered structure simulated probability form may the formation of fiber structure are relatively low, it can be inferred that the nucleation size fragments have more than four chain. At the same time, the simulation was found in the curly single beta sheet In the subsequent study, we found that these structures exist as a transition state that forms a larger, more ordered structure.
2. in the fourth chapter, for the simulation of inspection in the early four poly obtained with fiber characteristics of basic modules of the two structures, namely, single and double folded flat beta beta folding is the possibility of further formation of the structure module, we simulate seven dimer system 83-89 fragments. The study found that with the increase of the system. The chain to increase the number of hydrophobic interactions to play a crucial role on the structure formation and stability. A single beta sheet does not exist, the corresponding parallel double beta fold ratio increased significantly, and the formation of hydrophobic stability between layers. From our simulation results that parallel double beta kg stack the structure is microglobulin fragment of 83-89 fiber module possible gather configuration. In the simulation process, we also found that the beta barrel structure from the beta channel structure formed by folding curl, in the bucket Heart close by the hydrophobic core, nonpolar amino acid side chain atoms. The aggregation process of barrel can be formed by two ways. One is a single curl beta sheet through a combination of free single stranded extension formed; the other is directly through the double beta turn into vertical folding. Further study the beta barrel structure can be found, it can be transformed into each other and vertical double beta folding structure, which can be transformed into parallel double structure. So we believe that beta barrel structure is intermediate state of oligomeric aggregation.
The main contents of the 3. chapter of the fifth is in the computing and simulation time permitting, combining the MD-OPEP method with high efficiency, 83-89 fragment 16 poly molecular dynamics simulation. The body of the 6 simulated in the existing system, and there is no direct formation of ordered structure module form. That sphere of space structure the variety, in the hydrophobic interaction in strong hydrogen bonding interactions, the system requires a longer time to form a stable and orderly accumulation of hydrogen bonds, thus forming a stable structure of fiber module.
At the end of 4., in the sixth chapter, summarizes the previous research for human microglobulin 83-89 fragments of different systems. By comparing these three different size of the system, we found a series of 83-89 fragments of oligomeric ordered structures, to form and maintain these configurations of main interaction types, structure of fiber module the precursor, intermediate and transition special aggregation nucleation process.

【学位授予单位】：复旦大学
【学位级别】：硕士
【学位授予年份】：2008
【分类号】：R341

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