Isothermal titration calorimetry and surface plasmon resonance allow quantifying substrate binding to different binding sites of Bacillus subtilis xylanase

Sven Cuyvers, Emmie Dornez, Maher Abou Hachem, Birte Svensson, Michael Hothorn, Joanne Chory, Jan A. Delcour, Christophe M. Courtin

    Research output: Contribution to journalJournal articleResearchpeer-review

    Abstract

    Isothermal titration calorimetry and surface plasmon resonance were tested for their ability to study substrate binding to the active site (AS) and to the secondary binding site (SBS) of Bacillus subtilis xylanase A separately. To this end, three enzyme variants were compared. The first was a catalytically incompetent enzyme that allows substrate binding to both the AS and SBS. In the second enzyme, binding to the SBS was impaired by site-directed mutagenesis, whereas in the third enzyme, the AS was blocked using a covalent inhibitor. Both techniques were able to show that AS and SBS have a similar binding affinity.
    Original languageEnglish
    JournalAnalytical Biochemistry
    Volume420
    Issue number1
    Pages (from-to)90-92
    ISSN0003-2697
    DOIs
    Publication statusPublished - 2012

    Keywords

    • Isothermal titration calorimetry
    • Surface plasmon resonance
    • Xylanase
    • Xylooligosaccharides
    • Mechanism-based inhibitor

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