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Inducing Native-like Folding of Intrinsically Disordered Proteins using Stabilized a-Helical Peptides

使用稳定的α-螺旋肽诱导本质无序蛋白质的类似天然折叠

基本信息

批准号：
9333090
负责人：
Nicholas Alan Sawyer
金额：
$ 5.67万
依托单位：
NEW YORK UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2016
资助国家：
美国
起止时间：
2016-08-01 至 2019-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9333090
关键词：
Affinity Binding Binding Proteins Biological Assay C-terminal Cells Circular Dichroism Complement Complex Computer Simulation Disease EP300 gene Evaluation Fluorescence Fluorescence Polarization Genes Genetic Transcription Goals HIF1A gene Hydrogen Bonding Hypoxia Impairment In Vitro Laboratories Luciferases Malignant Neoplasms Measurement Measures Mediating Molecular Conformation Mus NMR Spectroscopy Peptides Play Protein Conformation Protein Family Proteins Quantitative Reverse Transcriptase PCR Research Role STAT2 gene Side Signal Transduction Specificity Structure TP53 gene Therapeutic Therapeutic Intervention Transcription Coactivator Transcriptional Activation Transcriptional Activation Domain Transcriptional Regulation Tumor Volume Up-Regulation Xenograft procedure alpha helix angiogenesis base biological systems covalent bond design experimental study flexibility functional group genome-wide hypoxia inducible factor 1 improved in vitro activity in vivo interest mimetics mimicry novel peptide structure peptidomimetics protein E protein aminoacid sequence protein folding protein function protein protein interaction protein structure small molecule tool tumor microenvironment tumor xenograft

项目摘要

PROJECT SUMMARY/ABSTRACT Intrinsically disordered proteins (IDPs) are a ubiquitous class of proteins whose structural plasticity allows them to function as hubs in protein-protein interaction (PPI) and signaling networks. IDP PPIs are therefore tightly regulated, and aberrant IDP PPIs are associated with many disease states. As IDP folding and activity are dependent on the conformations induced by its native binding partners, small molecules that induce native-like IDP conformations are highly desirable for studying IDP function. However, complex IDP PPI interfaces are difficult targets for traditional small molecules. Instead, non-traditional approaches using stabilized α-helical peptides, such as the hydrogen bond surrogate (HBS) approach developed in the Arora laboratory, are promising for creating small molecules that nucleate native-like IDP folding with both high affinity and high specificity. Thus, I hypothesize that HBS peptides can mimic key portions of IDP binding partners to nucleate IDP folding and competitively inhibit IDP PPIs in vitro and in vivo. The overall goal is to create HBS-derived artificial folders that induce IDPs to favor one fold over others. A classic IDP PPI example is the interaction of the general transcriptional coactivator p300 with the intrinsically disordered C-terminal transcription-activation domain of hypoxia-inducible factor 1 α (HIF1α CTAD). This interaction is critical for hypoxia-inducible transcription, leading to the upregulation of many cancer-associated genes involved in angiogenesis, invasion, and proliferation. The Arora lab has shown that mimicry of a HIF1α CTAD helical fragment downregulates hypoxia-inducible transcription and decreases xenograft tumor size in mice. However, targeting p300 may impair its ability to interact with its numerous binding partners. In the current proposal, I propose to develop a complementary strategy to use HBS peptides to induce HIF1α CTAD folding and competitively inhibit the p300-HIF1α CTAD interaction. To achieve this goal, I will first design and synthesize HBS peptides that mimic a p300 helical fragment that interacts with HIF1α CTAD. I will evaluate HBS peptide binding affinity and specificity for HIF1α CTAD in vitro. I will then characterize the structures of HIF1α CTAD-HBS complexes to evaluate HBS peptide-induced folding of HIF1α CTAD. I will also evaluate HBS peptides’ ability to modulate HIF1α-mediated transcription using cell-based assays. HBS peptides that induce native-like HIF1α CTAD folding with high affinity and specificity may serve as useful tools for studying HIF1α function and potential cancer therapeutic leads for HIF1α inhibition. This strategy for inducing IDP folding may also provide a specific application of a general approach to modulate IDP structures and activities using structured peptides.

项目总结/文摘