Site-wise Diversification of Combinatorial Libraries Using Insights from Structure-guided Stability Calculations.
Computational
Library design
Protein engineering
Site-wise diversification
Stability
Journal
Methods in molecular biology (Clifton, N.J.)
ISSN: 1940-6029
Titre abrégé: Methods Mol Biol
Pays: United States
ID NLM: 9214969
Informations de publication
Date de publication:
2022
2022
Historique:
entrez:
28
4
2022
pubmed:
29
4
2022
medline:
3
5
2022
Statut:
ppublish
Résumé
Many auspicious clinical and industrial accomplishments have improved the human condition by means of protein engineering. Despite these achievements, our incomplete understanding of the sequence-structure-function relationship prevents rapid innovation. To tackle this problem, we must develop and integrate new and existing technologies. To date, directed evolution and rational design have dominated as protein engineering principles. Even so, prior to screening for novel or improved functions, a large collection of variants, within a protein library, exist along an ambiguous mutational terrain. Complicating things further, the choice of where to initialize investigation along a vast sequence space becomes even more difficult given that the majority of any sequence lacks function entirely. Unfortunately, even when considering functionally relevant positions, random substitutions can prove to be destabilizing, causing a hindrance to an otherwise function-inducing, stability-reliant folding process. To enhance productivity in the field, we seek to address this issue of destabilization, and subsequent disfunction, at protein-protein and protein-ligand interacting regions. Herein, the process of choosing amenable positions - and amino acids at those positions - allows for a refined, knowledge-based approach to combinatorial library design. Using structural data, we perform computational stability prediction with FoldX's PositionScan and Rosetta's ddG_monomer in tandem, allowing for the refinement of our thermodynamic stability data through the comparison of results. In turn, we provide a process for selecting in silico predicted mutually stabilizing positions and avoiding overly destabilizing ones that guides the site-wise diversification of combinatorial libraries.
Identifiants
pubmed: 35482184
doi: 10.1007/978-1-0716-2285-8_3
doi:
Substances chimiques
Ligands
0
Proteins
0
Types de publication
Journal Article
Langues
eng
Sous-ensembles de citation
IM
Pagination
63-73Informations de copyright
© 2022. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.
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