Eldar Kim, Willy Wriggers, Martin Phillips, Kevin Kokabi, Peter Rubenstein, and Emil Reisler. Cross-Linking Constraints on F-actin Structure. J. Molec. Biol., 2000, Vol. 299, pp. 421-429.
The DNase I binding loop (residues 38-52), the hydrophobic plug (residues 262-274), and the C-terminus region are among the structural elements of monomeric (G-) actin that were proposed to form the intermonomer interface in F-actin. To test the proximity and interactions of these elements and to provide constraints on models of F-actin structure, cysteines were introduced into yeast actin either at residue 41 or 265. These mutations allowed for specific cross-linking of F-actin between C41 and C265, C265 and C374, and C41 and C265 using dibromobimane and disulfide bond formation. The cross-linked products were visualized on SDS PAGE and by electron microscopy. Model calculations carried out for the cross-linked F-actins revealed that considerable flexibility or displacement of actin residues is required in the disulfide cross-linked segments to fit these filaments into model F-actin structures. The calculated cross-linked structures showed a better fit to the Holmes than the refined Lorenz model of F-actin. It is predicted on the basis of such calculations that image reconstruction of electron micrographs of disulfide cross-linked C41-C374 F-actin should provide a conclusive test of these two similar models of F-actin structure.
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