ReBaTSA: A simplified CeTSA protocol for studying recombinant mutant proteins in bacterial extracts

ReBaTSA: A simplified CeTSA protocol for studying recombinant mutant proteins in bacterial extracts

The study of protein stability is crucial to biochemistry and relies on different methodologies. Recently, the Cellular Thermal Shift Assay has been introduced to study protein stability in whole cells. We report a novel application of CeTSA named ReBaTSA. This Recombinant Bacterial TSA was performe...

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Veröffentlicht in:Biochimica et biophysica acta. General subjects 2024-02, Vol.1868 (2), p.130526-130526, Article 130526
Hauptverfasser: Monticelli, Maria, Wright, Demi Marie, Cubellis, Maria Vittoria, Andreotti, Giuseppina
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container_title Biochimica et biophysica acta. General subjects
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creator Monticelli, Maria
Wright, Demi Marie
Cubellis, Maria Vittoria
Andreotti, Giuseppina
description The study of protein stability is crucial to biochemistry and relies on different methodologies. Recently, the Cellular Thermal Shift Assay has been introduced to study protein stability in whole cells. We report a novel application of CeTSA named ReBaTSA. This Recombinant Bacterial TSA was performed using clear extracts from bacteria expressing a recombinant protein, incubated at different temperatures, centrifuged and analyzed via SDS-PAGE. We demonstrated the feasibility and reliability of this simplified approach. We validated the method using the protein phosphomannomutase-2 and its common mutants, which were compared in the presence or the absence of a known ligand.
doi_str_mv 10.1016/j.bbagen.2023.130526
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title ReBaTSA: A simplified CeTSA protocol for studying recombinant mutant proteins in bacterial extracts
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