Conformational dynamics of loop 262-274 in G- and F-actin

Alexander Shvetsov, John D. Stamm, Martin Phillips, Dora Warshaviak, Christian Altenbach, Peter A. Rubenstein, Kálmán Hideg, Wayne L. Hubbell, Emil Reisler

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According to the original Holmes model of F-actin structure, the hydrophobic loop 262-274 stabilizes the actin filament by inserting into a pocket formed at the interface between two protomers on the opposing strand. Using a yeast actin triple mutant, L180C/L269C/C374A [(LC)2CA], we showed previously that locking the hydrophobic loop to the G-actin surface by a disulfide bridge prevents filament formation. We report here that the hydrophobic loop is mobile in F- as well as in G-actin, fluctuating between the extended and parked conformations. Copper-catalyzed, brief air oxidation of (LC)2CA F-actin on electron microscopy grids resulted in the severing of thin filaments and their conversion to amorphous aggregates. Disulfide, bis(methanethiosulfonate) (MTS), and dibromobimane (DBB) cross-linking reactions proceeded in solution at a faster rate with G- than with F-actin. Cross-linking of C180 to C269 by DBB (4.4 Å) in either G- or F-actin resulted in shorter and less stable filaments. The cross-linking with a longer MTS-6 reagent (9.6 Å) did not impair actin polymerization or filament structure. Myosin subfragment 1 (S1) and tropomyosin inhibited the disulfide cross-linking of phalloidin-stabilized F-actin. Electron paramagnetic resonance measurements with nitroxide spin-labeled actin revealed strong spin-spin coupling and a similar mean interspin distance (∼10 Å) in G- and in F-actin, with a broader distance distribution in G-actin. These results show loop 262-274 fluctuations in G- and F-actin and correlate loop dynamics with actin filament formation and stability.

Original languageEnglish
Pages (from-to)6541-6549
Number of pages9
Issue number20
Publication statusPublished - May 23 2006


ASJC Scopus subject areas

  • Biochemistry

Cite this

Shvetsov, A., Stamm, J. D., Phillips, M., Warshaviak, D., Altenbach, C., Rubenstein, P. A., Hideg, K., Hubbell, W. L., & Reisler, E. (2006). Conformational dynamics of loop 262-274 in G- and F-actin. Biochemistry, 45(20), 6541-6549.