NEBULIN, A HELICAL ACTIN-BINDING PROTEIN

Nebulin, a giant protein (molecular mass 800 kDa) specific for the ske letal muscle of vertebrates, has been suggested to be involved in the length regulation of the thin filament as a 'molecular ruler'. Despite its size, nebulin appears to be composed mainly: of small repeats of similar to 35 amino acids. We have characterized in this study the con formational and functional properties of single repeats. Complete repe ats were found to bind to F-actin while a truncated one did not. One r epeat is therefore the smallest unit for nebulin-actin interaction. Ci rcular dichroism and nuclear magnetic resonance spectra measured for t he peptides in water indicated a transient helical conformation. The f olded region is located for them all around the conserved sequence SDx xYK. The helical conformation is strongly stabilized by anionic deterg ents and trifluoroethanol while uncharged or positively charged deterg ents have no effect. Since the surface of the actin filament is known to contain clusters of negative charges, anionic detergents may mimic the effect of an actin environment. 3D structures were calculated for three representative peptides in SDS. In vivo, the nebulin helices sho uld form a complex with the actin filament. Based on the assumed impor tance of charge interactions between nebulin and actin, we propose a m odel for the structure of the F-actin-nebulin complex in vivo. Accordi ng to that, two nebulin molecules occupy symmetrical positions along t he central cleft of the actin filament bridging the two strands of the actin two-start helix. The consistency of this model with experimenta l data is discussed.