The effects of ADF/cofilin and profilin on the conformation of the ATP-binding cleft of monomeric actin

Roland Kardos, Kinga Pozsonyi, Elisa Nevalainen, Pekka Lappalainen, Miklos Nyitrai, Gabor Hild

    Research output: Contribution to journalArticleScientificpeer-review

    Abstract

    Actin depolymerizing factor (ADF)/cofilin and profilin are small actin-binding proteins, which have central roles in cytoskeletal dynamics in all eukaryotes. When bound to an actin monomer, ADF/cofilins inhibit the nucleotide exchange, whereas most profilins accelerate the nucleotide exchange on actin monomers. In this study the effects of ADF/cofilin and profilin on the accessibility of the actin monomer's ATP-binding pocket was investigated by a fluorescence spectroscopic method. The fluorescence of the actin bound epsilon-ATP was quenched with a neutral quencher (acrylamide) in steady-state and time dependent experiments, and the data were analyzed with a complex form of the Stern-Volmer equation. The experiments revealed that in the presence of ADF/cofilin the accessibility of the bound epsilon-ATP decreased, indicating a closed and more compact ATP-binding pocket induced by the binding of ADF/cofilin. In the presence of profilin the accessibility of the bound e-ATP increased, indicating a more open and approachable protein matrix around the ATP-binding pocket. The results of the fluorescence quenching experiments support a structural mechanism regarding the regulation of the nucleotide exchange on actin monomers by ADF/cofilin and profilin.
    Original languageEnglish
    JournalBiophysical Journal
    Volume96
    Issue number6
    Pages (from-to)2335-2343
    Number of pages9
    ISSN0006-3495
    DOIs
    Publication statusPublished - 2009
    MoE publication typeA1 Journal article-refereed

    Fields of Science

    • 1182 Biochemistry, cell and molecular biology

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