Protein Binder (ProBi) as a New Class of Structurally Robust Non-Antibody Protein Scaffold for Directed Evolution
| Autor: | Štěpán Herynek, Lucie Kolářová, Gustavo Fuertes, Jiří Černý, Phuong Ngoc Pham, Bohdan Schneider, Petr Kolenko, Pavel Mikulecky, Jiří Zahradník, Jiří Pavlíček, Tatsiana Charnavets, Lada Biedermannová, Maroš Huličiak |
|---|---|
| Jazyk: | angličtina |
| Rok vydání: | 2021 |
| Předmět: |
0301 basic medicine
Scaffold protein Scaffold Protein Conformation interleukin-10 lcsh:QR1-502 Computational biology lcsh:Microbiology Article 03 medical and health sciences 0302 clinical medicine Protein structure computational saturation Virology protein scaffold Computer Simulation Amino Acid Sequence directed evolution Databases Protein Thermostability Chemistry Protein Stability ribosome display Proteins protein engineering computer.file_format Protein engineering Protein Data Bank Directed evolution Recombinant Proteins 030104 developmental biology Infectious Diseases Ribosome display Directed Molecular Evolution computer Ribosomes 030217 neurology & neurosurgery Protein Binding |
| Zdroj: | Viruses Volume 13 Issue 2 Viruses, Vol 13, Iss 190, p 190 (2021) |
| ISSN: | 1999-4915 |
| DOI: | 10.3390/v13020190 |
| Popis: | Engineered small non-antibody protein scaffolds are a promising alternative to antibodies and are especially attractive for use in protein therapeutics and diagnostics. The advantages include smaller size and a more robust, single-domain structural framework with a defined binding surface amenable to mutation. This calls for a more systematic approach in designing new scaffolds suitable for use in one or more methods of directed evolution. We hereby describe a process based on an analysis of protein structures from the Protein Data Bank and their experimental examination. The candidate protein scaffolds were subjected to a thorough screening including computational evaluation of the mutability, and experimental determination of their expression yield in E. coli, solubility, and thermostability. In the next step, we examined several variants of the candidate scaffolds including their wild types and alanine mutants. We proved the applicability of this systematic procedure by selecting a monomeric single-domain human protein with a fold different from previously known scaffolds. The newly developed scaffold, called ProBi (Protein Binder), contains two independently mutable surface patches. We demonstrated its functionality by training it as a binder against human interleukin-10, a medically important cytokine. The procedure yielded scaffold-related variants with nanomolar affinity. |
| Databáze: | OpenAIRE |
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