Genetically encoded bicyclic peptide libraries for the discoveryof novel antiviral agents

用于发现新型抗病毒药物的基因编码双环肽库

基本信息

批准号：
10189880
负责人：
Jeffery Micheal Tharp
金额：
$ 9.52万
依托单位：
YALE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2021
资助国家：
美国
起止时间：
2021-09-02 至 2022-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10189880
关键词：
2019-nCoV Address Adoption Affinity Amines Amino Acids Amino Acyl-tRNA Synthetases Antibodies Antiviral Agents Bacteriophage M13 Bacteriophages Binding Cells Chemicals Communicable Diseases Coronavirus Coronavirus Infections Coupled Cyclic Peptides Cyclization Cysteine Development Development Plans Diagnosis Diagnostic Directed Molecular Evolution Disease Drug Targeting Engineering Evolution Future Genetic Code Goals Human In Vitro Laboratories Libraries Ligands Lung diseases Mediating Membrane Fusion Mentors Methods Middle East Respiratory Syndrome Coronavirus Modeling Molecular Conformation Peptide Conformation Peptide Library Peptides Phage Display Phase Prevention Property Proteins Reaction Research Ribosomes Route SARS coronavirus Side Supervision Surface System Techniques Technology Therapeutic Training Transfer RNA Translations Tyrosine-tRNA Ligase Virus Work base career career development combinatorial functional group human coronavirus in vivo inhibitor/antagonist novel novel coronavirus novel therapeutics pandemic disease peptide drug prevent programs protein protein interaction scaffold screening skills small molecule technology development thioether tool

项目摘要

PROJECT SUMMARY Bicyclic peptides are conformationally constrained peptides comprised of two macrocyclic rings. Owing to their increased rigidity, bicyclic peptides can bind to protein targets with greater affinity and selectivity than their linear and monocyclic counterparts. As a result, these molecules are highly desirable scaffolds for the development of peptide-based therapeutics. Phage display is a laboratory evolution technique that enables the discovery of high- affinity peptide ligands from large, combinatorial peptide libraries. Although initially limited to linear peptides, this technique has been adapted for the discovery of bicyclic peptide ligands. Most often, phage-displayed bicyclic peptides are generated by chemically modifying linear peptides using cysteine-reactive small molecules; how- ever, this method is technically challenging. As a result, progress in this field has been limited. Recently, several studies have used genetic code expansion to co-translationally install cysteine-reactive noncanonical amino acids (ncAAs) into phage-displayed peptides to produce libraries of cyclic peptides. This strategy has significant advantages over the chemical cyclization approach, but is currently limited to monocyclic peptides. The over- arching objective of this proposal is to develop technology that enables phage display of bicyclic pep- tides using genetic code expansion. Our central hypothesis is that bifunctional ncAAs, i.e. ncAAs containing two cysteine-reactive functional groups, can be used to generate ribosomally synthesized bicyclic peptides by intramolecular reaction with cysteine residues. To realize our objective, we will pursue three Specific Aims. In Aim 1 (K99 Phase) we will engineer an aminoacyl-tRNA synthetase that recognizes bifunctional ncAAs contain- ing two cysteine-reactive moieties. This will be accomplished using traditional and state-of-the-art methods of directed evolution. In Aim 2 (K99/R00 Phase) we will develop a phage display system that is compatible with co- translational installation of bifunctional ncAAs and we will optimize this system for bicyclic peptide formation. We will then validate this system by selecting and characterizing ligands for model targets. In Aim 3 (R00 Phase) we will use phage display to identify bicyclic peptides that bind to the spike protein of human coronaviruses and inhibit virus-host membrane fusion. By targeting proteins from various coronaviruses, we will strive to identify inhibitors with broad-spectrum antiviral activity. The proposed work will provide a facile route for generating bi- cyclic peptide libraries thereby greatly accelerating the discovery of therapeutic peptide leads. The Candidate, Dr. Jeffery Tharp’s long-term career goal is to establish an independent research program that uses genetic code expansion and phage display to develop antiviral peptides for the diagnosis, treatment, and prevention of infectious diseases. Herein we propose a detailed five-year Career Development Plan supervised by mentors Drs. Dieter Söll and Craig Wilen, and a team of subject-matter experts. This plan will augment previous training and address key training gaps to prepare Dr. Tharp for accomplishing his long-term career goal.

项目摘要自行车宠物是由两个大环环组成的构象约束宠物。由于他们的刚性提高，生物petides可以与线性更高的亲和力和选择性结合蛋白质靶标和单核对应物。结果，这些分子是高度可取的支架基于肽的治疗。噬菌体显示是一种实验室进化技术，可以发现高 - 来自大型组合肽库的亲和肽配体。尽管最初仅限于线性肽，但技术已适应生物肽配体的发现。最常见的是噬菌体播种的生物学肽是通过使用半胱氨酸反应性小分子对线性修饰宠物产生的。如何- 曾经，这种方法在技术上受到挑战。结果，该领域的进展受到限制。最近，几个研究已使用遗传密码扩展来同时安装半胱氨酸反应性非有可能的氨基酸（NCAA）成噬菌体斑点肽，以产生环状肽的文库。该策略具有重要的优于化学环化方法的优势，但目前仅限于单核肽。超过该提案的拱门目标是开发能够噬菌体展示生物学的技术潮汐使用遗传代码扩展。我们的中心假设是双功能NCAA，即含有NCAA 两个半胱氨酸反应性官能团可用于通过与半胱氨酸保留的分子内反应。为了实现我们的目标，我们将追求三个具体目标。 AIM 1（K99阶段）我们将设计一种氨基酰基-TRNA合成酶，该酶识别含有双功能的NCAAS- 为两个半胱氨酸反应性部分。这将使用传统和最先进的方法来完成定向进化。在AIM 2（K99/R00相）中，我们将开发一个与共同兼容的噬菌体显示系统双功能NCAA的翻译安装，我们将优化该系统以形成生物肽。我们然后，将通过选择和表征模型目标的配体来验证该系统。在AIM 3（R00阶段）中我们将使用噬菌体显示器来识别与人冠状病毒和抑制病毒宿主膜融合。通过靶向来自各种冠状病毒的蛋白质，我们将努力识别具有广谱抗病毒活性的抑制剂。拟议的工作将提供一条轻松的途径，以产生双因此，环状肽库很大程度上加速了热肽导管的发现。候选人， Jeffery Tharp博士的长期职业目标是建立一个独立的研究计划，该计划使用通用代码扩展和噬菌体显示，以开发抗病毒胡椒剂，以诊断，治疗和预防传染病。在此，我们提出了一项详细的五年职业发展计划，由导师监督博士。 DieterSöll和Craig Wilen，以及主题专家团队。该计划将增加以前的培训并解决关键培训差距，以准备萨尔普博士完成他的长期职业目标。