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Cryo-electron microscopy structures of pyrene-labeled ADP-Pi- and ADP-actin filaments

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  • Steven Z. Chou

    (Yale University, PO Box 208103)

  • Thomas D. Pollard

    (Yale University, PO Box 208103
    Yale University, PO Box 208103
    Yale University, PO Box 208103)

Abstract

Since the fluorescent reagent N-(1-pyrene)iodoacetamide was first used to label skeletal muscle actin in 1981, the pyrene-labeled actin has become the most widely employed tool to measure the kinetics of actin polymerization and the interaction between actin and actin-binding proteins. Here we report high-resolution cryo-electron microscopy structures of actin filaments with N-1-pyrene conjugated to cysteine 374 and either ADP (3.2 Å) or ADP-phosphate (3.0 Å) in the active site. Polymerization buries pyrene in a hydrophobic cavity between subunits along the long-pitch helix with only minor differences in conformation compared with native actin filaments. These structures explain how polymerization increases the fluorescence 20-fold, how myosin and cofilin binding to filaments reduces the fluorescence, and how profilin binding to actin monomers increases the fluorescence.

Suggested Citation

  • Steven Z. Chou & Thomas D. Pollard, 2020. "Cryo-electron microscopy structures of pyrene-labeled ADP-Pi- and ADP-actin filaments," Nature Communications, Nature, vol. 11(1), pages 1-9, December.
    • Handle: RePEc:nat:natcom:v:11:y:2020:i:1:d:10.1038_s41467-020-19762-1
    DOI: 10.1038/s41467-020-19762-1
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