Role of the Dictyostelium 30 kDa protein in actin bundle formation.

We have studied the formation of bundles in mixtures of actin with the Dictyostelium 30 kDa actin-bundling protein as a function of 30 kDa protein concentration, actin concentration, and filament length. The presence of the 30 kDa protein promotes formation of filament bundles at actin concentrations and filament lengths that are not spontaneously aligned into liquid crystalline domains in the absence of the 30 kDa protein. Bundle formation in the presence of the 30 kDa protein was observed over a broad range of actin filament lengths and concentrations. Bundling was filament length dependent, and short filaments were more efficiently bundled. Bundles formed at actin concentrations as low as 2 microM. The volume fraction of the bundled portion and concentrations of actin and the 30 kDa protein in the bundled portion were measured using a sedimentation assay. Bundles have concentrations of actin and 30 kDa protein that are 10-20 and 5-20 times, respectively, greater than that of the bulk solution. Computer modeling reveals that bundling of actin by a bundling protein increases both the mean length and the polydispersity of the length distribution, factors which lower the actin concentration required for spontaneous alignment within the bundle. We propose that entropy-driven spontaneous ordering may contribute to bundle formation in two ways. Bundling of actin creates longer aggregates with a more polydisperse length distribution in which actin aligns spontaneously within the bundle at very low concentrations. In addition, bundling creates locally high concentrations of actin within these aggregates that will spontaneously align, providing an additional driving force for bundle ordering.