Fluorescence measurements of Ca2+ binding to domain VI of calpain

J. Simon C. Arthur, John S. Elce

    Research output: Chapter in Book/Report/Conference proceedingChapter (peer-reviewed)

    Abstract

    The Ca2+ binding properties of calpain are of great interest, both biochemically in the wider context of EF-hand proteins, and physiologically, in the context of calpain regulation. There are two major parameters which one might wish to measure: the actual number of binding sites (n) and the binding constants for Ca2+. The latter is normally the macroscopic binding constant for Ca2+ of the molecule as a whole (Kd), or the microscopic binding constants for each of the EF-hands, but for cooperative binding these are much more difficult to measure. There is evidence of various kinds to suggest that Ca2+ binding causes conformational change in the whole molecule, and this forms the basis for measuring Ca2+ binding by means of changes in fluorescence.

    Original languageEnglish
    Title of host publicationCalpain Methods and Protocols
    EditorsJohn S. Elce
    Place of PublicationTotowa, New Jersey
    PublisherHumana Press
    Pages121-127
    Number of pages7
    EditionPart II
    ISBN (Electronic)9781592590506
    ISBN (Print)9780896036321
    DOIs
    Publication statusPublished - 2000

    Publication series

    NameMethods in Molecular Biology
    PublisherHumana Press
    Volume144
    ISSN (Print)1064-3745
    ISSN (Electronic)1940-6029

    Keywords

    • Rats
    • Animals
    • Calcium
    • Spectrometry, Fluorescence
    • Recombinant Proteins
    • Kinetics
    • Calpain
    • Protein Structure, Tertiary
    • Protein Structure, Quaternary
    • Binding Sites

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