Publication Details

FireProt 2.0: Web-based Platform for the Fully-Automated Design of Thermostable Proteins

MUSIL Miloš, JEŽÍK Andrej, HORÁČKOVÁ Jana, BORKO Simeon, KABOUREK Petr, DAMBORSKÝ Jiří and BEDNÁŘ David. FireProt 2.0: Web-based Platform for the Fully-Automated Design of Thermostable Proteins. Briefings in Bioinformatics, vol. 25, no. 1, 2023, pp. 1-10. ISSN 1467-5463.
Czech title
FireProt 2.0: Webová platforma pro plně automatizovaný návrh stabilních proteinů
journal article
Musil Miloš, Ing., Ph.D. (DIFS FIT BUT)
Ježík Andrej, Bc. (FIT BUT)
Horáčková Jana, Mgr. (LL)
Borko Simeon, Ing. (FIT BUT)
Kabourek Petr (LL)
Damborský Jiří, prof. Mgr., Dr. (LL)
Bednář David, Mgr. (LL)

ancestral, back-to-consensus, B-factor, epistasis, evolution, force-field, multiple-point mutant, protein engineering, saturation mutagenesis, thermostability


Thermostable proteins find their use in numerous biomedical and biotechnological applications. However, the computational design of stable proteins often results in single-point mutations with a limited effect on protein stability. However, the construction of stable multiple-point mutants can prove difficult due to the possibility of antagonistic effects between individual mutations. FireProt protocol enables the automated computational design of highly stable multiple-point mutants. FireProt 2.0 builds on top of the previously published FireProt web, retaining the original functionality and expanding it with several new stabilization strategies. FireProt 2.0 integrates the AlphaFold database and the homology modelling for structure prediction, enabling calculations starting from a sequence. Multiple-point designs are constructed using the Bron-Kerbosch algorithm minimizing the antagonistic effect between the individual mutations. Users can newly limit the FireProt calculation to a set of user-defined mutations, run a saturation mutagenesis of the whole protein, or select rigidifying mutations based on B-factors. Evolution-based back-to-consensus strategy is complemented by ancestral sequence reconstruction. FireProt 2.0 is significantly faster and a reworked graphical user interface broadens the tool's availability even to users with older hardware. FireProt 2.0 is freely available at

Briefings in Bioinformatics, vol. 25, no. 1, ISSN 1467-5463
Oxford University Press
EID Scopus
   author = "Milo\v{s} Musil and Andrej Je\v{z}\'{i}k and Jana Hor\'{a}\v{c}kov\'{a} and Simeon Borko and Petr Kabourek and Ji\v{r}\'{i} Damborsk\'{y} and David Bedn\'{a}\v{r}",
   title = "FireProt 2.0: Web-based Platform for the Fully-Automated Design of Thermostable Proteins",
   pages = "1--10",
   journal = "Briefings in Bioinformatics",
   volume = 25,
   number = 1,
   year = 2023,
   ISSN = "1467-5463",
   doi = "10.1093/bib/bbad425",
   language = "english",
   url = ""
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