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Substrate targeting mechanism of a DesCEND pathway

DesCEND 通路的底物靶向机制

基本信息

批准号：
10624332
负责人：
Bing Hao
金额：
$ 42.17万
依托单位：
UNIVERSITY OF CONNECTICUT SCH OF MED/DNT
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-08-15 至 2024-05-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10624332
关键词：
Address Affinity Apoptotic Binding Biochemical Bioinformatics Biological Assay C-terminal Calorimetry Calpain Cell Culture System Cell physiology Cells Co-Immunoprecipitations Complement Complex Cyclin D1 DNA Damage Disease Drug Targeting Elements Endopeptidases F Box Domain F-Box Proteins Fluorescence Polarization G1 Arrest Genotoxic Stress Goals Human Imaging Techniques In Vitro Knowledge Libraries MAP2K6 gene Maps Mass Spectrum Analysis Measurement Measures Mediating Methods Molecular Mutagenesis Mutation N-terminal Pathogenesis Pathway interactions Peptides Play Printing Property Proteins Proteolysis Research Roentgen Rays Role Signal Transduction Site Snails Specificity Stimulus Structure Substrate Specificity Surface System Tail Testing Therapeutic Titrations Ubiquitin Ubiquitination Work biophysical analysis biophysical properties candidate selection design experimental study improved in vitro Assay insight intermolecular interaction live cell imaging mutant novel protein aminoacid sequence protein degradation proteostasis rational design reconstitution recruit response structural determinants treatment strategy ubiquitin-protein ligase

项目摘要

Project Summary Ubiquitin-mediated protein degradation is essential to maintaining proteostasis, thereby controlling diverse cellular processes. The specificity of protein ubiquitination is largely determined by recognition of short peptide motifs known as degrons in protein substrates by E3 ubiquitin ligases. F-box proteins serve as substrate- recognition components of the Skp1–Cul1–F-box-protein (SCF) E3 superfamily. The F-box protein FBXO31 mediates cyclin D1 degradation to induce G1 arrest after DNA damage. The C-terminal region of cyclin D1 contains sequences rich in Pro, Glu/Asp, Ser and Thr. This so-called PEST motif is enriched in highly dynamic disordered regions of eukaryotic proteins and serves as degradation signal for many unstable proteins. We have recently determined the X-ray crystal structure of a C-terminal PEST-containing cyclin D1 peptide bound to Skp1–FBXO31. The structure reveals that specific recognition is achieved by an intricate inter-molecular interaction network involving as well the cyclin D1 carboxyl tail that occupies an open surface cavity in the - barrel motif of FBXO31. We show that this PEST degron mediates the interaction of cyclin D1 with FBXO31 for ubiquitination in vitro and proteolysis in cells. We hypothesize that FBXO31 is responsible for recognition of C- terminal PEST-containing substrate degrons through a novel DesCEND (destruction via C-end degrons) pathway. By identifying and characterizing substrates of FBXO31 at both a systems level and the molecular level, we aim to elucidate the complementary structural determinants of the FBXO31–degron pair that confer substrate specificity and dictate protein recognition and fate. To achieve our goals, we have developed the following specific aims to: 1) identify and characterize C-end substrate degrons targeted by FBXO31 in human proteins; 2) characterize the structural and biochemical properties of candidate FBXO31 degrons; and 3) characterize endoproteolytic cleavage-generated FBXO31 degrons.

项目总结