Illuminating Notch receptor-ligand selectivity through structure-guided protein engineering

通过结构引导的蛋白质工程阐明Notch受体-配体选择性

• 批准号: 10727696
• 负责人: Vincent Christopher Luca
• 金额: $ 2.26万
• 依托单位: H. LEE MOFFITT CANCER CTR & RES INST
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-09-01 至 2024-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10727696
• 关键词: Affinity; Architecture; Area; Binding; Biological; Biology; Biophysics; Cell Communication; Cell Surface Receptors; Cells; Chemicals; Complex; Cues; Development; Directed Molecular Evolution; Discrimination; Electron Microscopy; Engineering; Environment; Funding; Goals; Kinetics; Laboratories; Language; Length; Ligands; Malignant Neoplasms; Mammals; Mediating; Molecular; Mutation; Outcome; Output; Pathway interactions; Post-Translational Protein Processing; Protein Engineering; Proteins; Research; Roentgen Rays; Series; Signal Transduction; Structure; Therapeutic; Tissues; Training; Vision; Visualization; Work; antagonist; aortic valve disorder; biophysical properties; cell type; design; experimental study; glycosylation; human disease; insight; interdisciplinary approach; mechanical force; mechanical properties; notch protein; novel; novel therapeutics; programs; receptor; reconstitution; response; structural biology; translational applications; transmission process

PROJECT SUMMARY Overall vision of the research program. My research is focused on understanding the “molecular language” of cell-to-cell communication. Cell surface receptors transmit signals in response to external cues, and receptor activity is regulated by extrinsic factors such as posttranslational modifications, mechanical force, and chemical environment. Using a multidisciplinary approach that leverages my training in structural biology, biophysics, and protein engineering, my group is attempting to solve difficult problems in the area of receptor biology. We hypothesize that the distinct structural and biophysical parameters governing receptor-ligand interactions control downstream signaling, and that ligands may be engineered to alter these parameters and reprogram signaling outputs. My long-term goal is to harness the insight obtained through our mechanistic studies to design proteins that can fine-tune receptor activity and function as powerful next-generation therapeutics. Overview of research in the laboratory. Notch signaling is initiated when a mechanosensory Notch receptor is “pulled on” by a Delta-like (DLL) or Jagged (Jag) ligand expressed on an adjacent cell. In mammals, the various Notch receptor (Notch1-4) and ligand (DLL1, DLL4, Jag1, Jag2) subtypes are able to induce distinct or even opposing cellular responses. Ligand selectivity is further regulated by Notch receptor glycosylation, which inhibits the activity of Jag ligands yet potentiates the activity of DLL ligands. We currently have a poor understanding of the molecular basis for ligand discrimination in the Notch pathway, and structural studies of Notch receptor-ligand interactions have been difficult because their nearly undetectable binding affinity precludes reconstitution of stable complexes. We recently overcame this obstacle by using directed evolution to strengthen DLL4 and Jag1 binding to Notch1, which allowed us to capture their interacting domains for x-ray crystal structure determination. My lab is now building upon these transformative results to explore how the structural, biophysical, and mechanical properties of various ligand subtypes contribute to their unique functions. Goals for the next five years. Over the five-year funding period, the goals of this project are to: (i) systematically characterize the structure, binding kinetics, and signaling potency of Notch receptor-ligand complexes; (ii) use electron microscopy to visualize the ultrastructural architecture of full-length Notch1-DLL4 and Notch1-Jag1 complexes; (iii) determine the biophysical basis by which Notch glycosylation regulates DLL and Jag activity, and (iv) design tissue-specific Notch antagonists. The successful completion of this work will establish the molecular mechanisms of Notch receptor-ligand selectivity and will guide the engineering of novel modulators of Notch signaling with expanded capabilities in a variety of biological and therapeutic contexts.

项目总结

项目成果

Soluble and multivalent Jag1 DNA origami nanopatterns activate Notch without pulling force.

• DOI: 10.1038/s41467-023-44059-4
• 发表时间: 2024-01-18
• 期刊: NATURE COMMUNICATIONS
• 影响因子: 16.6
• 作者: Smyrlaki, Ioanna;Foerdos, Ferenc;Rocamonde-Lago, Iris;Wang, Yang;Shen, Boxuan;Lentini, Antonio;Luca, Vincent C.;Reinius, Bjoern;Teixeira, Ana I.;Hoegberg, Bjoern
• 通讯作者: Hoegberg, Bjoern