The fibronectin type III domain as a scaffold for novel binding proteins
Open accessAbstract
The fibronectin type III domain (FN3) is a small autonomous folding unit which occurs in many animal proteins involving in ligand binding. The beta-sandwich structure of FN3 closely resembles that of immunoglobulin domains. We have prepared a phage display library of FN3 in which residues in two surface loops were randomized. We have selected mutant FN3s which bind to a test ligand, ubiquitin, with significant affinities, while the wild-type FN3 shows no measurable affinity. A dominant clone was expressed as a soluble protein and its properties were investigated in detail. Heteronuclear NMR characterization revealed that the selected mutant protein retains the global fold of FN3. It also has a modest conformational stability despite mutations at 12 out of 94 residues. These results clearly show the potential of FN3 as a scaffold for engineering novel binding proteins.
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Monomeric proteins based on the synthetic protein (FN3) skeleton are similar in function to antibodies and can be used in combination with the corresponding target; they have smaller molecular weight and clearer structure compared to traditional antibodies, and are more stable in oxidation/reduction conditions, making them a new type of ideal inhibitor.
“Monomeric proteins, based on the synthetic protein (FN3) skeleton, are similar in function to antibodies and can be used in combination with the corresponding target. These proteins have a smaller molecular weight and clearer structure compared to traditional antibodies. Moreover, they are more stable in oxidation/reduction conditions, making them a new type of ideal inhibitor.330,331”
From Recent advances in targeting the “undruggable” proteins: from drug discovery to clinical trials (2023) · cites this paper as [330]
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 [110]
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References 51 · Cited by 585
Research works printed but not linked: 3
- Kunkel, T. A. (1985). Rapid and ef®cient site-speci®c mutagenesis without phenotypic selection. Proc. Natl Acad. Sci. USA, 82, 488± 492.
- Rosenblum, J. S. & Barbas, C. F. I. (1995). Synthetic anti- bodies. In Antibody Engineering (Borrenbaeck, C. A. K., ed.), pp. 89 ± 116, Oxford University Press, Oxford.
- Sambrook, J., Fritsch, E. F. & Maniatis, T. (1989). Molecu- lar Cloning: A Laboratory Manual, 2nd edit., Cold Spring Harbor Laboratory Press, Cold Spring Har- bor, NY.
- Authors
- Koide, A., Bailey, C. W., Huang, X., & Koide, S.
- Affiliations
- University of Rochester Medicine (US)
- Corresponding author
- Koide, Shohei
- Venue
- Journal of Molecular Biology
- Publisher
- Elsevier BV
- Published
- 1998-12
- Volume
- 284
- Issue
- 4
- Pages
- 1141-1151
- ISSN
- 0022-2836 (ISSN-L)
- OpenAlex
- W2024298562
- Cited by
- 585
- References
- 51
- Citations by year
- 2026: 15 · 2025: 25 · 2024: 32 · 2023: 33 · 2022: 28 · 2021: 35 · 2020: 34 · 2019: 21 · 2018: 28 · 2017: 32 · 2016: 27 · 2015: 30 · 2014: 21 · 2013: 25 · 2012: 23
- FWCI
- 1.51
- Type
- journal article
- Language
- en
- Open access
- hybrid
- doi.org
- Licence
- Locations
- Journal of Molecular Biology (publishedVersion, cc-by-nc-nd)
- PubMed (publishedVersion)
- Funders
- National Institutes of Health (5r29gm055042-05)
- National Institute of General Medical Sciences (GM55042)
- Topics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Protein Structure and Dynamics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Biochemical and Structural Characterization
- Health Sciences › Medicine › Radiology, Nuclear Medicine and Imaging › Monoclonal and Polyclonal Antibodies Research
- Keywords
- fibronectin type III domain, protein scaffolds, phage display, ubiquitin binding, beta-sandwich structure, immunoglobulin-like domains, conformational stability, ligand binding
- MeSH terms
- Amino Acid Sequence, Animals, Base Sequence, Binding Sites, Carrier Proteins, DNA, Recombinant, Fibronectins, Humans, Models, Molecular, Molecular Sequence Data, Magnetic Resonance Spectroscopy, Protein Conformation, Recombinant Proteins, Protein Engineering, Mutagenesis, Bacteriophage M13, Protein Folding, In Vitro Techniques
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