Continuous evolution of compact protein degradation tags regulated by selective molecular glues

• 成果类型: Article
• 署名作者: Mercer, Jaron A. M.; Decarlo, Stephan J.; Burman, Shourya S. Roy; Sreekanth, Vedagopuram; Nelson, Andrew T.; Hunkeler, Moritz; Chen, Peter J.; Donovan, Katherine A.; Kokkonda, Praveen; Tiwari, Praveen K.; Shoba, Veronika M.; Deb, Arghya; Choudhary, Amit; Fischer, Eric S.; Liu, David R.
• 署名单位: Harvard University; Massachusetts Institute of Technology (MIT); Broad Institute; Massachusetts Institute of Technology (MIT); Harvard University; Harvard University; Howard Hughes Medical Institute; Harvard University; Harvard University Medical Affiliates; Dana-Farber Cancer Institute; Harvard University; Harvard Medical School; Harvard University; Massachusetts Institute of Technology (MIT); Broad Institute; Harvard University; Harvard Medical School; Harvard University; Harvard University Medical Affiliates; Brigham & Women's Hospital
• 刊物名称: SCIENCE
• ISSN/ISSBN: 0036-8990
• DOI: 10.1126/science.adk4422
• 发表日期: 2024-03-15
• 关键词: continuous directed evolution; phage-assisted evolution; cryo-em; escherichia-coli; human-cells; MODEL; identification; orientation; activation; stringency

摘要

Conditional protein degradation tags (degrons) are usually >100 amino acids long or are triggered by small molecules with substantial off-target effects, thwarting their use as specific modulators of endogenous protein levels. We developed a phage-assisted continuous evolution platform for molecular glue complexes (MG-PACE) and evolved a 36-amino acid zinc finger (ZF) degron (SD40) that binds the ubiquitin ligase substrate receptor cereblon in complex with PT-179, an orthogonal thalidomide derivative. Endogenous proteins tagged in-frame with SD40 using prime editing are degraded by otherwise inert PT-179. Cryo-electron microscopy structures of SD40 in complex with ligand-bound cereblon revealed mechanistic insights into the molecular basis of SD40's activity and specificity. Our efforts establish a system for continuous evolution of molecular glue complexes and provide ZF tags that overcome shortcomings associated with existing degrons.