Accurate de novo design of high-affinity protein-binding macrocycles using deep learning.

Journal: Nature chemical biology
Published Date: Jun 20, 2025

Abstract

Developing macrocyclic binders to therapeutic proteins typically relies on large-scale screening methods that are resource intensive and provide little control over binding mode. Despite progress in protein design, there are currently no robust approaches for de novo design of protein-binding macrocycles. Here we introduce RFpeptides, a denoising diffusion-based pipeline for designing macrocyclic binders against protein targets of interest. We tested 20 or fewer designed macrocycles against each of four diverse proteins and obtained binders with medium to high affinity against all targets. For one of the targets, Rhombotarget A (RbtA), we designed a high-affinity binder (K < 10 nM) despite starting from the predicted target structure. X-ray structures for macrocycle-bound myeloid cell leukemia 1, γ-aminobutyric acid type A receptor-associated protein and RbtA complexes match closely with the computational models, with a Cα root-mean-square deviation < 1.5 Å to the design models. RFpeptides provides a framework for rapid and custom design of macrocyclic peptides for diagnostic and therapeutic applications.

Authors

  • Stephen A Rettie
    Molecular and Cellular Biology program, University of Washington, Seattle, WA, USA.
  • David Juergens
    Department of Biochemistry, University of Washington, Seattle, WA 98105.
  • Victor Adebomi
    Institute for Protein Design, University of Washington, Seattle, WA, USA.
  • Yensi Flores Bueso
    Institute for Protein Design, University of Washington, Seattle, WA, USA.
  • Qinqin Zhao
    Department of Microbiology, University of Washington, Seattle, WA, USA.
  • Alexandria N Leveille
    Department of Chemistry, Tufts University, 62 Talbot Avenue, Medford, MA, USA.
  • Andi Liu
    Center for Precision Health, McWilliams School of Biomedical Informatics, The University of Texas Health Science Center at Houston, Houston, TX, USA.
  • Asim K Bera
    Department of Biochemistry, University of Washington, Seattle, WA, USA.
  • Joana A Wilms
    Heinrich-Heine-Universität Düsseldorf, Institut für Physikalische Biologie, Düsseldorf, Germany.
  • Alina Üffing
    Heinrich-Heine-Universität Düsseldorf, Institut für Physikalische Biologie, Düsseldorf, Germany.
  • Alex Kang
    Department of Biochemistry, University of Washington, Seattle, WA, USA.
  • Evans Brackenbrough
    Institute for Protein Design, University of Washington, Seattle, WA 98105, USA.
  • Mila Lamb
    Department of Biochemistry, University of Washington, Seattle, WA, USA.
  • Stacey R Gerben
    Department of Biochemistry, University of Washington, Seattle, WA, USA.
  • Analisa Murray
    Department of Biochemistry, University of Washington, Seattle, WA, USA.
  • Paul M Levine
    Department of Biochemistry, University of Washington, Seattle, WA, USA.
  • Maika Schneider
    Department of Medicinal Chemistry, University of Washington, Seattle, WA, USA.
  • Vibha Vasireddy
    Department of Medicinal Chemistry, University of Washington, Seattle, WA, USA.
  • Sergey Ovchinnikov
    Center for Systems Biology, Harvard University, Cambridge, MA 02138, United States.
  • Oliver H Weiergräber
    Forschungszentrum Jülich, Institute of Biological Information Processing, Structural Biochemistry (IBI-7), Jülich, Germany.
  • Dieter Willbold
    Heinrich-Heine-Universität Düsseldorf, Institut für Physikalische Biologie, Düsseldorf, Germany.
  • Joshua A Kritzer
    Department of Chemistry, Tufts University, 62 Talbot Avenue, Medford, MA, USA.
  • Joseph D Mougous
    Department of Microbiology, University of Washington, Seattle, WA, USA.
  • David Baker
    Department of Biochemistry, University of Washington, Seattle, Washington.
  • Frank DiMaio
    Department of Biochemistry, University of Washington, Seattle, WA 98195, USA.
  • Gaurav Bhardwaj
    Discipline of Ophthalmology and Eye Health and Save Sight Institute, Sydney Medical School, Faculty of Medicine and Health, The University of Sydney, Sydney, Australia.

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