Database Commons

a catalog of worldwide biological databases

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 modeling 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 http://loschmidt.chemi.muni.cz/fireprotweb.

The majority of naturally occurring proteins have evolved to function under mild conditions inside the living organisms. One of the critical obstacles for the use of proteins in biotechnological applications is their insufficient stability at elevated temperatures or in the presence of salts. Since experimental screening for stabilizing mutations is typically laborious and expensive, in silico predictors are often used for narrowing down the mutational landscape. The recent advances in machine learning and artificial intelligence further facilitate the development of such computational tools. However, the accuracy of these predictors strongly depends on the quality and amount of data used for training and testing, which have often been reported as the current bottleneck of the approach. To address this problem, we present a novel database of experimental thermostability data for single-point mutants FireProtDB. The database combines the published datasets, data extracted manually from the recent literature, and the data collected in our laboratory. Its user interface is designed to facilitate both types of the expected use: (i) the interactive explorations of individual entries on the level of a protein or mutation and (ii) the construction of highly customized and machine learning-friendly datasets using advanced searching and filtering. The database is freely available at https://loschmidt.chemi.muni.cz/fireprotdb.