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PubMed · 11026684

Intermolecular dynamics and function in actin filaments.

E Kim·E Reisler
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Abstract

Structural models of F-actin suggest that three segments in actin, the DNase I binding loop (residues 38-52), the hydrophobic plug (residues 262-274) and the C-terminus, contribute to the formation of an intermolecular interface between three monomers in F-actin. To test these predictions and also to assess the dynamic properties of intermolecular contacts in F-actin, Cys-374 pyrene-labeled skeletal alpha-actin and pyrene-labeled yeast actin mutants, with Gln-41 or Ser-265 replaced with cysteine, were used in fluorescence experiments. Large differences in Cys-374 pyrene fluorescence among copolymers of subtilisin-cleaved (between Met-47 and Gly-48) and uncleaved alpha-actin showed both intra- and intermolecular interactions between the C-terminus and loop 38-52 in F-actin. Excimer band formation due to intermolecular stacking of pyrene probes attached to Cys-41 and Cys-265, and Cys-41 and Cys-374, in mutant yeast F-actin confirmed the proximity of these residues on the paired sites (to within 18 A) in accordance with the models of F-actin structure. The dynamic properties of the intermolecular interface in F-actin formed by loop 38-52, plug 262-274 and the C-terminus may account for the observed cross-linking of these sites with reagents < 18 A. The functional importance of actin filament dynamics was demonstrated by the inhibition of the in vitro motility in the Gln-41-Cys-374 cross-linked actin filaments.

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Actin CytoskeletonActinsAmino Acid SubstitutionAnimalsCross-Linking ReagentsElectrophoresis, Polyacrylamide GelFluorescent DyesFungal ProteinsModels, MolecularMutagenesis, Site-DirectedProtein ConformationPyrenesSpectrometry, FluorescenceJournal ArticleResearch Support, U.S. Gov't, Non-P.H.S.Research Support, U.S. Gov't, P.H.S.

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E Kim, E Reisler. 2000-08-30. Intermolecular dynamics and function in actin filaments.. https://doi.org/10.1016/s0301-4622(00)00143-5

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