Electrostatic Roles on Macromolecular Assemblies in Vision Processes

看东西的能力是人类和他/她的世界的接口。这种能力经常受到大分子及其组装的故障的阻碍，这些大分子和组装构成了人眼的几个隔室。例子包括人体晶状体中的晶体蛋白和人眼玻璃体中的透明质酸-胶原蛋白复合凝胶。尽管了解这些带电大分子在人眼中的行为具有重大意义，但缺乏基本的理解。挑战来自于晶体蛋白序列的变化，以及它们如何通过疏水和静电相互作用的组合相互作用。以玻璃体为例，胶原蛋白分子如何在透明质酸基质形成的限制性凝胶状环境中组织成纤维，是定量和基础研究的新领域。首席研究员建议使用合成的组合，包括定点诱变和化学合成，计算机建模，使用光散射，显微镜和流变学的表征技术，以及先进的聚合物物理理论。拟议研究的一个主要组成部分是为下一代科学家提供指导和建立科学基础，以便他们在一个对消除人类失明至关重要的领域进行基础研究。技术摘要：为了了解与人体晶状体相关的晶体蛋白的聚集/分解，以及与人体玻璃体相关的透明质酸基质中胶原束的组装，提出了实验和建模，使用定点诱变，化学合成，静态和动态光散射，差分动态显微镜，流变学和计算机建模。具体来说，该提案解决了(i)晶体蛋白序列（通过定点突变合成）在聚集中的作用，（ii）晶体蛋白的电荷修饰在聚集/分解中的作用，（iii）胶原束的成核和生长，以及（iv）胶原束在透明质酸基质中的异质分布程度。提出的工作旨在确定在水拥挤环境中大量蛋白质分子聚集背后的基本原理，基于构象波动和静电力的可控调制。提出的体外实验将对人眼所需的光透明度的大分子起源有深入的了解。培训学生在这个跨学科领域，重点是基本理解是拟议活动的强大教育组成部分。该奖项反映了美国国家科学基金会的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准进行评估，被认为值得支持。

NON-TECHNICAL SUMMARYThe ability to see is the interface between a human and the rest of his/her universe. This ability is often hampered by malfunctioning of macromolecules and their assemblies which make up several compartments of human eye. Examples include crystallin proteins in human lens and hyaluronic acid-collagen composite gel in the vitreous of human eye. Despite the enormous significance of knowing how these charged macromolecules behave in the human eye, a fundamental understanding is lacking. The challenge arises from variations in the sequences of the crystallin proteins and how they interact among themselves via a combination of hydrophobic and electrostatic interactions. In the case of vitreous, how the collagen molecules organize into fibers inside a restrictive gel-like environment created by hyaluronic acid matrix, is a new area for quantitative and fundamental investigation. The Principal Investigator proposes to use a combination of synthesis, including site-directed mutagenesis and chemical synthesis, computer modeling, characterization techniques using light scattering, microscopy, and rheology, and advanced polymer physics theories. One of the major components of the proposed research is to mentor and build scientific foundation for the next generation of scientists to pursue fundamental research in an area of tremendous importance in getting rid of human blindness.TECHNICAL SUMMARYExperiments and modeling are proposed to understand the aggregation/disaggregation of crystallin proteins pertinent to human lens, and the assembly of collagen bundles in hyaluronic acid matrixes pertinent to human vitreous, using site-directed mutageneis, chemical synthesis, static and dynamic light scattering, differential dynamic microscopy, rheology, and computer modeling. Specifically, the proposal addresses (i) role of crystallin protein sequences (synthesized by site-directed mutagenesis) on aggregation, (ii) role of charge modification of crystallin proteins on aggregation/disaggregation, (iii) nucleation and growth of collagen bundles, and (iv) extent of heterogeneous distribution of collagen bundles inside hyaluronic acid matrix. The proposed work aims at identifying fundamental principles behind aggregation of large numbers of protein molecules in aqueous crowded environments, based on conformational fluctuations and controllable modulations of electrostatic forces. The proposed in vitro experiments will result in insights on the macromolecular origins for required transparency to light in human eye. Training students in this interdisciplinary area with a strong focus on fundamental understanding is the strong educational component of the proposed activity.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.