Fast model systems for misfolding, binding and aggregation

用于错误折叠、绑定和聚合的快速模型系统

• 批准号: 8727044
• 负责人: MARTIN GRUEBELE
• 金额: $ 26.97万
• 依托单位: UNIVERSITY OF ILLINOIS AT URBANA-CHAMPAIGN
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-05-01 至 2017-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8727044
• 关键词: Adult; Amyloid; Amyloidosis; Binding; Binding Proteins; Binding Sites; Biological Models; Chimera organism; Communities; Complement; Complex; Computers; Coupled; Data; Deglutition; Development; Device or Instrument Development; Diffusion; Disease; Dissociation; Elements; Engineering; Equilibrium; Event; Experimental Models; Eyelid structure; Feedback; Free Energy; Future; Generations; Goals; Grant; Illinois; Kinetics; Knowledge; Lasers; Learning; Measures; Micelles; Mining; Modeling; Mutation; Nucleic Acids; Pathway interactions; Plant Roots; Population; Postdoctoral Fellow; Process; Proteins; RNA; RNA Binding; RNA-Binding Proteins; Reaction; Relaxation; Relaxation Techniques; Research; Resolution; Signal Transduction; Site; Solutions; Sorting - Cell Movement; Speed; Structure; Students; System; Technology; Time; U1A protein; United States National Institutes of Health; Visit; Work; base; crosslink; improved; instrument; interest; intermolecular interaction; meetings; millisecond; molecular dynamics; monomer; next generation; protein aggregation; protein folding; prototype; public health relevance; research study; simulation; single molecule; success; supercomputer; temperature jump

DESCRIPTION (provided by applicant): Fast (microsecond) experiments and molecular dynamics simulation are finally working hand-in-hand to provide validated atomistic pictures of protein folding dynamics. Larger proteins and millisecond folders are around the corner, although empirical force fields need more improvement before mechanistic predictions become as reliable as average rate coefficients or native structures. It is time to extend this simulation experiment interplay to misfolding, aggregation and binding processes. The problem is that such processes are usually slow - seconds to days. In order to compare experiment and simulation directly during the next 4 years, when simulations will not reach far beyond the 1 ms regime yet, the solution is simple: create small and fast experimental model systems for misfolding, aggregation and binding. These are analogous to fast two-state and downhill folding studied during the last 10 years: certainly not all proteins fold that way, but much useful was learned from our ability to directly compare such model proteins with simulation. Here, we propose development of: 1) ?6-85 as a model system for sheet-containing misfolded traps (T denaturation and T-jumps) as well as excess helix-containing traps (P denaturation, P-jumps). The key is that all folding and misfolding events are completed in ~ 1 ms or faster. 2a) Fast-folding tethered proteins (WW and ?6-85) to facilitate the interplay between transient aggregation and folding. Oligomeric aggregates include chimeras with elements swapped among monomers, as well as less structured aggregates. Tethering allows high effective concentrations without going to high protein concentration (leading to uncontrollable aggregation), and also helps keep MD simulation by keeping reactants in close proximity. 2b) U1A-RNA binding to multiple sites on U1A protein will be a fast model system for binding at multiple sites. 3) For the next generation of misfolding/aggregation/binding research, we will develop a single molecule instrument capable of high throughput (106 molecules/day) and studying 2 or more molecules interacting without cross-linking them or confining them in a micelle. Five simulation groups have been brought on board (Schulten, Shaw, Cheung, Luthey-Schulten and Pande), and their students/postdocs are already working closely with mine so simulation can be developed in parallel with the experiments. The goal is to provide data in the few microsecond to few millisecond range, amenable to full atom simulation over the next 4 years, so misfolding/aggregation/binding processes can be studied at the atomistic level, but on systems small and fast enough for simulation to work. We build on our knowledge of ?6-85, WW domain and U1A, so much is known experimentally and computationally about the monomeric building blocks of our misfolding/aggregation/binding models.

描述(由申请人提供)：FAST(微秒)实验和分子动力学模拟终于携手合作，提供了经过验证的蛋白质折叠动力学的原子图像。更大的蛋白质和毫秒文件夹即将到来，尽管经验力场需要更多的改进，才能使机械预测变得像平均速率系数或自然结构一样可靠。现在是扩展这种模拟的时候了 实验相互作用于错误折叠、聚集和结合过程。问题是，这样的过程通常很慢，只有几秒钟到几天。为了在接下来的4年里直接比较实验和模拟，当模拟还不会远远超过1ms的时候，解决方案很简单：创建用于错误折叠、聚集和结合的小而快速的实验模型系统。这些类似于过去10年中研究的快速两态和下坡折叠：当然，并不是所有的蛋白质都以这种方式折叠，但从我们直接将此类模型蛋白质与模拟进行比较的能力中学到了很多有用的东西。在这里，我们建议发展：1)？6-85作为含有片状错误折叠的陷阱(T变性和T-跳跃)以及含有过量螺旋的陷阱(P变性，P-跳跃)的模型系统。关键是所有折叠和错误折叠事件都在~1ms或更快的时间内完成。2)快速折叠系留蛋白(WW和？6-85)，以促进瞬时聚集和折叠之间的相互作用。寡聚聚集体包括单体之间交换元件的嵌合体，以及结构较差的聚集体。系留允许高有效浓度而不会达到高蛋白质浓度(导致无法控制的聚集)，并通过保持反应物的紧密接近来帮助保持MD模拟。2B)与U1a蛋白上多个位点结合的U1a-RNA将成为多个位点结合的快速模型系统。3)对于下一代错误折叠/聚集/结合的研究，我们将开发一台高通量(106个分子/天)的单分子仪器，研究两个或更多分子之间的相互作用，而不需要交联剂或将它们限制在胶束中。五个模拟小组(Schulten、Shaw、Cheung、Luhe-Schulten和Pande)已经加入，他们的学生/博士后已经与我的团队密切合作，因此模拟可以与实验并行开发。我们的目标是提供几微秒到几毫秒范围内的数据，在接下来的4年里可以进行全原子模拟，这样就可以在原子水平上研究错误折叠/聚集/结合过程，但要在足够小和足够快的系统上进行模拟。我们建立在我们对？6-85、WW结构域和U1a的知识基础上，所以很多关于我们错误折叠/聚集/结合模型的单体构件的实验和计算都是已知的。