The kinetics of actin polymerization were analyzed by taking into account nucleation, elongation, and spontaneous fragmentation of filaments. Polymerization curves measured in the presence of potassium (40 mM) were found to be in good agreement with curves calculated for the assumption that nucleation and elongation but no fragmentation reactions occur. Polymerization curves measured in the presence of calcium (1.8 mM) or magnesium (0.6 mM MgCl2 and 0.5 mM EGTA) could only be stimulated by calculated curves when spontaneous fragmentation was assumed to occur in addition to nucleation and elongation. The experiments reported in this study that even in the absence of ultrasonication or shear forces actin filaments may break spontaneously and that the extent of fragmentation depends strongly on the experimental conditions. Spontaneous fragmentation changes the shape of the polymerization curves significantly. When fragmentation of filaments takes place, a relatively long lag phase of polymerization is observed that is followed by a strongly increasing polymerization rate to reach the final constant value quickly. On the other hand, when filaments are formed exclusively by nucleation, the polymerization curves approach the final constant value slowly after a relatively short initial lag phase.

Dielectric constant and loss of the membrane-active peptide alamethicin in octanol/dioxane mixtures have been measured at frequencies between 5 kHz and 50 MHz. On the basis of a rotational mechanism of dipolar orientation, the observed dispersion provides information regarding size, shape, and dipole moment of the structural entities which the solute may assume in media of diverse lipophilicity. Particularly detailed results are obtained in a pure octanol solvent where an apparent molecular weight of alamethicin could be determined. It turns out that in this quite lipophilic medium most of the peptide material exists as a monomer particle that has approximate length and diameter of 35 and 13 A, respectively. It carries a dipole moment of approximately 75 Debye units (directed nearly parallel to the long axis). At our concentrations of a few milligrams per milliliters, appreciable formation of dimers by head-to-tail linkage is indicated. When the octanol content is reduced by adding greater amounts of dioxane, larger particles are encountered. This is accompanied by a decrease of the effective polarity. The inherent increase of hydrophilicity in the dioxane-enriched solvent apparently favors another monomer conformation that has a low dipole moment and easily aggregates to some kind of micelle.

No abstract

Dielectric constant and loss of alamethicin in solvents of various degrees of lipophilicity (namely mixtures of n-octanol and dioxane) have been measured at frequencies from 5 kHz to 50 MHz. By means of a conventional Cole-Cole approach, dielectric properties were evaluated to obtain information about the structural and dipolar properties of the peptide in view of its function as a voltage-dependent pore former in membranes. The results for a pure octanol solvent (together with an apparent molecular weight determined by ultracentrifugation) indicate the existence of primarily monomeric particles of quite elongated shape and of comparatively high dipole moment. Adding dioxane was found to yield considerable aggregation and a decrease of polarity.