Self-Assembly of Collagen-like Octadecamers Based on the Stem Region of C1q and Related Proteins

在化学系的大分子、超分子和纳米化学项目的支持下，杰弗里·D。威廉·马什·赖斯大学的哈特杰林克正在研究 在结构上模仿胶原蛋白的小肽的自组装。 胶原蛋白是人体中最丰富的蛋白质，在伤口愈合、组织再生、癌症转移和软硬组织的整体完整性中起着重要作用。 这种蛋白质由三条多肽链组成，它们以紧密的三螺旋缠绕在一起。 每条多肽链由氨基酸序列组成，这些氨基酸序列通过称为肽键或连接的特殊化学键连接。 当计算所有氨基酸时，胶原蛋白是分子量接近300，000 g/mol的非常大的大分子。 这种尺寸限制了对有关胶原蛋白中三螺旋如何在溶液中组装和形成的原子水平信息的获取。 为了解决这个问题，研究小组将专注于胶原蛋白中两个不同区域的较小片段，即六个三重螺旋捆绑在一起的“茎”和三重螺旋彼此向外张开形成独立三重螺旋的“分支”。 这两个区域的多肽链将被合成，并使用一套复杂的化学技术进行彻底的表征。 将利用计算算法SCEPTTr来辅助实验设计，并确定自组装所需的最小氨基酸序列。 分支区域的不同序列具有提供良好折叠的三螺旋系统的潜力，这将扩大对三螺旋稳定性的理解。 另一方面，茎区可以提供确定平行堆积三重螺旋的原子水平结构的机会。 该研究将为本科和研究生教育课程提供出色的基础，因为它是多学科的性质，并结合了化学设计和合成与传统的结构生物学和医学应用。 外展计划将招募高中学生和他们的教师在服务不足的社区参加研究在夏季和开发令人兴奋的课程，整合这项工作。 SCEPTTr将得到增强，并继续分发给社区，旨在帮助设计更大的肽模拟物。 本研究将集中在基于C1 q干区的胶原样十八聚体和相关防御胶原的自组装。 在第一个目标中，将制备具有取自防御胶原蛋白分支区域的序列的胶原蛋白模拟肽（CMP）。这些化学上不同的序列将被选择用于自组装研究，因为它们显示出折叠的趋势。 将使用标准Fmoc化学通过固相肽合成制备肽，并表征其热稳定性。 还将独立检查关键的成对相互作用。 该稳定性数据将用于改善SCEPTTr准确建模的序列范围。 生物衍生的序列将通过共价捕获来稳定，并测试其结合防御胶原蛋白的已知配体的能力。 在最后一个目标中，将合成具有取自防御胶原的茎区的序列的CMP并使其寡聚化以形成十八聚体。 将进行序列研究，以评估十八聚体自组装所需的最小区域。 将通过NMR和X射线晶体学进一步表征良好折叠的十八聚体。螺旋束将通过共价捕获来稳定，并测试其结合相关配体的能力。这个项目，如果成功的话，将提高对胶原蛋白的多步骤，分层自组装的一般理解，并提供潜在的生物医学应用的CMP的评估。 除了从这种结构研究中获得的基本生物物理知识外，防御胶原蛋白在先天免疫应答以及炎症和免疫的整体协调中发挥的关键作用将为制备新的肽创造许多机会，该奖项反映了NSF的法定使命，并通过使用基金会的智力价值和更广泛的评估被认为值得支持。影响审查标准。

With the support of the Macromolecular, Supramolecular and Nanochemistry program in the Division of Chemistry, Professor Jeffrey D. Hartgerink of William Marsh Rice University is studying self-assembly of small popypeptides that structurally mimic collagen. Collagen is the most abundant protein in the human body and plays major roles in wound healing, tissue regeneration, cancer metastasis and the overall integrity of both soft and hard tissues. This protein is composed of three polypeptide chains that are wrapped together in a tight triple helix. Each polypeptide chain consists of sequences of amino acids that are linked by a particular kind of chemical bond called a peptide bond or link. When counting all amino acids, collagen is a very large macromolecule with molecular weight approaching 300,000 g/mol. This size limits the access to atomic level information about how the triple helix in collagen is assembled and formed in solution. To circumvent this problem, the research team will focus on smaller fragments from two different regions in collagen, namely “stem” where the six triple helices bundle together and the “branches” where the triple helices splay outward from one another making independent triple helices. Polypeptides chains from these two regions will be synthesized and thoroughly characterized using a suite of sophisticated chemical techniques. A computational algorithm SCEPTTr will be utilized to aid in the experimental design and identify the minimal sequences of amino acids needed for self-assembly. Diverse sequences of the branch region have the potential to provide well folded triple helical systems that will expand understanding of triple helix stabilization. The stem region, on the other hand, could offer opportunity to determine the atomic level structure of parallel packed triple helices. The research will provide an outstanding basis for undergraduate and graduate educational programs because it is multi-disciplinary in nature and combines chemical design and synthesis with traditional structural biology and medical application. Outreach programs will recruit high school students and their teachers in underserved communities to participate in research over the summer and develop exciting curriculum that integrates this work. SCEPTTr will be enhanced and continued to be distributed to the community with the aim to assist in the design of larger peptide mimics. This research will focus on the self-assembly of collagen-like octadecamers based on the stem region of C1q and related Defense Collagens. In the first objective, Collagen Mimetic Peptides (CMPs) with sequences taken from the branch region of Defense Collagens will be prepared. These chemically diverse sequences will be selected for self-assembly studies because they show a tendency to fold. Peptides will be prepared by solid phase peptide synthesis using standard Fmoc chemistry and characterized for their thermal stability. Key pairwise interactions will also be independently examined. This stability data will be used to improve the range of sequences accurately modeled by SCEPTTr. Biologically derived sequences will be stabilized through covalent capture and tested for their ability to bind known ligands of Defense Collagens. In the last objective, CMPs with sequences taken from the stem region of Defense Collagens will be synthesized and oligomerized to form octadecamers. A sequence study will be performed to assess the minimal regions required for octadecamer self-assembly. Well folded octadecamers will be further characterized by NMR and X-ray crystallography. Helical bundles will be stabilized by covalent capture and tested for their ability to bind relevant ligands. This project, if successful, will improve general understanding of the multi-step, hierarchical self-assembly of collagen and provide an assessment of CMPs for potential biomedical applications. In addition to the fundamental biophysical knowledge gained from this structural investigation, the critical role that Defense Collagens play in the innate immune response and the overall orchestration of inflammation and immunity will create many opportunities for preparing novel peptide-based drugs and biopolymers.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.