Cited by 656Monoclonal and Polyclonal Antibodies ResearchBiochemical and Structural CharacterizationChemical Synthesis and Analysis

High-affinity binders selected from designed ankyrin repeat protein libraries

Binz, H Kaspar, Amstutz, Patrick, Kohl, Andreas, Stumpp, Michael T, Briand, Christophe, Forrer, Patrik, Grütter, Markus G, Plückthun, Andreas·2004·Nature Biotechnology

Abstract

We report here the evolution of ankyrin repeat (AR) proteins in vitro for specific, high-affinity target binding. Using a consensus design strategy, we generated combinatorial libraries of AR proteins of varying repeat numbers with diversified binding surfaces. Libraries of two and three repeats, flanked by 'capping repeats,' were used in ribosome-display selections against maltose binding protein (MBP) and two eukaryotic kinases. We rapidly enriched target-specific binders with affinities in the low nanomolar range and determined the crystal structure of one of the selected AR proteins in complex with MBP at 2.3 A resolution. The interaction relies on the randomized positions of the designed AR protein and is comparable to natural, heterodimeric protein-protein interactions. Thus, our AR protein libraries are valuable sources for binding molecules and, because of the very favorable biophysical properties of the designed AR proteins, an attractive alternative to antibody libraries.

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Crossref · 10.1038/nbt962

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  • POI-targeting polypeptides can either be single chain antibodies (e.g. nanobodies) or a variety of synthetic polypeptide scaffolds that bind POIs such as monobodies, DARPINs, affimers, or Affibodies.

    “These POI-targeting polypeptides can either be single chain antibodies (e.g. variable domains from camelid or shark-derived heavy chain only antibodies, nanobodies), or a variety of synthetic polypeptide scaffolds that bind POIs such as fibronectin type III domain-based monobodies [110, 111], designed ankyrin repeats (DARPINs) [112, 113], cystatin-derived affimers [114] or Affibodies derived from”

    From Advances in targeted degradation of endogenous proteins (2019) · cites this paper as [113]

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Authors
Binz, H. K., Amstutz, P., Kohl, A., Stumpp, M. T., Briand, C., Forrer, P., Grütter, M. G., & Plückthun, A.
Affiliations
University of Zurich (CH)
Corresponding author
  • Grütter, Markus G
  • Plückthun, Andreas
Venue
Nature Biotechnology
Published
2004-04-18
Volume
22
Issue
5
Pages
575-582
ISSN
  • 0733-222X (ISSN-L)
  • 1087-0156
  • 1546-1696
OpenAlex
W2084938664
Cited by
656
References
51
Citations by year
2026: 7 · 2025: 22 · 2024: 17 · 2023: 26 · 2022: 27 · 2021: 23 · 2020: 31 · 2019: 24 · 2018: 22 · 2017: 38 · 2016: 28 · 2015: 36 · 2014: 33 · 2013: 33 · 2012: 35
FWCI
30.73
Type
journal article
Language
en
Open access
closed
Locations
  • Nature Biotechnology (publishedVersion)
  • PubMed (publishedVersion)
Funders
  • Bundesministerium für Bildung und Forschung
  • National Center of Competence in Research Chemical Biology
Topics
  • Health Sciences › Medicine › Radiology, Nuclear Medicine and Imaging › Monoclonal and Polyclonal Antibodies Research
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Biochemical and Structural Characterization
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Chemical Synthesis and Analysis
Keywords
ankyrin repeat proteins, combinatorial protein libraries, ribosome display, protein-protein interaction, maltose-binding protein, high-affinity binders, consensus design, protein crystal structure
MeSH terms
Amino Acid Sequence, Enzyme-Linked Immunosorbent Assay, Molecular Sequence Data, Protein Conformation, Ankyrin Repeat

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