Fundamental Processes in the Folding of Helical Proteins

蛋白质折叠，即最初未折叠的蛋白质快速自我组装成折叠功能形式的过程，通常被称为遗传密码的后半部分。蛋白质折叠所需的所有信息都编码在一级序列中，但仍然缺乏对折叠过程的详细了解。对蛋白质折叠的兴趣持续增长，部分原因是人们认识到折叠的基本和实际重要性，部分原因是实验和理论的进步。计算模拟和实验之间的协同作用的潜力产生了相当大的兴趣，折叠非常迅速的小蛋白质。这些都是解决蛋白质折叠中的基本问题和开发新方法的绝佳测试案例。该项目旨在推断α-螺旋蛋白质折叠的一般机制特征，测试关于其折叠的特定假设，开发研究其折叠的新方法，并为计算研究提供严格的基准。一个关键目标是将实验结果与折叠的计算研究联系起来。为此，研究主要集中在绒毛蛋白头段螺旋子域（HP-36）。HP-36是最小的天然存在的协同折叠序列，是已知折叠最快的蛋白质之一。它的快速折叠，小尺寸和简单的结构使其成为蛋白质折叠的计算和理论研究中非常受欢迎的模型系统。 本研究要解决的关键问题包括：（1）未折叠态系综的本质是什么？它在快速折叠中的作用是什么？(2)蛋白质折叠的分子动力学模拟到底有多精确，它们提供了折叠途径的可靠图像还是仅仅是天然状态的模型？(3)折叠速率限制步骤的本质是什么？将使用肽模型、不稳定突变体、pH依赖性稳定性测量和模拟来研究未折叠状态系综。新开发的同位素编辑IR检测T-跳跃方法将用于研究野生型HP-36和改变未折叠状态的突变体的折叠。对蛋白质折叠的进一步理解对基础生物学以及生物技术和生物材料设计具有重要的实际意义。 蛋白质折叠问题的解决方案在于化学，生物学和物理学的接口，研究将涉及实验和理论小组之间的密切合作。这些因素为参与研究生和本科生创造了一个刺激的多学科研究环境。 斯托尼布鲁克是美国种族最多样化的主要研究型大学之一，这项研究将为传统上代表性不足的群体增加机会。本科生参与是研究的一个关键组成部分。与本科院校建立了伙伴关系，以扩大本科生的参与超越斯托尼布鲁克。一个“采用实验室”计划正在开发作为第一年本科荣誉实验室课程的一部分，以提供广泛的学生与现代研究的初步接触。传统的本科生实验练习正在修改，以更好地将其与现代科学联系起来。例如，某些食用染料是蛋白质聚集的抑制剂。聚集可以使用廉价的本科分光计进行浊度测量进行监测。该实验说明了光散射，蛋白质结构，抑制的概念，并提供了一个介绍蛋白质折叠，同时很好地结合到目前的研究。实验室练习也将作为斯托尼布鲁克的大学范围内的计划，旨在提供支持和鼓励对科学和工程感兴趣的本科妇女的一部分。

Protein folding, the processes by which an initially unfolded protein rapidly self assembles into its folded functional form, has often been called the second half of the genetic code. All of the information required for a protein to fold is encoded in the primary sequence but a detailed understanding of the folding process is still lacking. Interest in protein folding continues to grow, driven in part by the realization of the fundamental and practical importance of folding and in part by experimental and theoretical advances. The potential for synergy between computational simulation and experiments has generated considerable interest in small proteins that fold very rapidly. These are excellent test cases for addressing fundamental issues in protein folding and for developing new methodology. This project aims to deduce general mechanistic features of the folding of alpha-helical proteins, to test specific hypotheses about their folding, to develop new methods to study their folding, and to provide rigorous bench marks for computational studies. A key goal is to link experimental results with computational studies of folding. To that end, the research is primarily focused on the villin headpiece helical subdomain (HP-36). HP-36 is the smallest naturally occurring sequence which folds cooperatively and it is one of the fastest folding proteins known. Its rapid folding, small size and simple structure have made it an extraordinarily popular model system for computational and theoretical studies of protein folding. Key questions to be addressed by this research include: (1) What is the nature of the unfolded state ensemble and what is its role in rapid folding? (2) Just how accurate are molecular dynamic simulations of protein folding, do they provide a reliable picture of the folding pathway or just a model of the native state? (3) What is the nature of the rate limiting step for folding? The unfolded state ensemble will be studied using peptide models, destabilized mutants, pH dependent stability measurements and simulations. Newly developed isotope edited IR detected T-jump methods will be used to study the folding of wild type HP-36 and mutants which alter the unfolded state. An improved understanding of protein folding has important implications for basic biology as well as important practical implications for biotechnology and for biomaterials design. The solution of the protein folding problem lies at the interface of chemistry, biology and physics and the research will involve close collaboration between experimental and theoretical groups. These factors create a stimulating multidisciplinary research environment for participating graduate and undergraduate students. Stony Brook is one of the most ethnically diverse major research universities in the nation and the research will enhance opportunities for traditionally underrepresented groups. Undergraduate participation is a key component of the research. Partnerships with undergraduate institutions have been established to expand undergraduate participation beyond Stony Brook. An "adopt a lab" program is being developed as part of the first year undergraduate honors lab class in order to provide a wide range of students with initial exposure to modern research. Traditional undergraduate laboratory exercises are being revised to better tie them into modern science. For example, certain food dyes are inhibitors of protein aggregation. Aggregation can be monitored using turbidity measurements conducted with inexpensive undergraduate spectrometers. The experiment illustrates light scattering, protein structure, the concept of inhibition and provides an introduction to protein folding while nicely tying into current research. Lab exercises will also be developed as part of Stony Brook's university-wide program designed to provide support and encouragement to undergraduate women who are interested in science and engineering.