High-affinity binders selected from designed ankyrin repeat protein libraries

@article{Binz2004HighaffinityBS,
  title={High-affinity binders selected from designed ankyrin repeat protein libraries},
  author={H. Kaspar Binz and Patrick Amstutz and Andreas Kohl and Michael T. Stumpp and Christophe Briand and Patrik Forrer and Markus G. Grütter and Andreas Pl{\"u}ckthun},
  journal={Nature Biotechnology},
  year={2004},
  volume={22},
  pages={575-582}
}
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… 
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611 Citations
Highly Influential Citations
16
Background Citations
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Methods Citations
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Results Citations
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611 Citations

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TLDR
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TLDR
The generation of a library built upon the framework of a consensus FN3 domain sequence resulting in binding proteins the authors call Centyrins is described, which provides important data contributing to the understanding of potential FN3 binding interfaces and a new tool for generating high-affinity scaffold molecules.
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TLDR
The use of designed ankyrin repeat proteins as tailor-made intracellular kinase inhibitors that bind APH with high specificity and with affinities down to the subnanomolar range is demonstrated.
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TLDR
This work describes biophysical and structural analyses required to extend existing binder scaffolds and simplifies an existing protocol for the assembly of highly diverse synthetic binder libraries.
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TLDR
The crystal structures of two complexes between αRep and their cognate targets were solved and show that the new interfaces are established by the variable surfaces of the repeated modules, as well as the variable N-cap residues.
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Destabilizing an interacting motif strengthens the association of a designed ankyrin repeat protein with tubulin
TLDR
Biochemical and structural characterizations demonstrated that the evolved mutants achieved a gain in affinity through destabilization of the C-cap, which relieves the need of a DARPin conformational change upon tubulin binding and removes unfavorable interactions in the complex.
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