Abstract
We used a cone and plate rheometer to evaluate the mechanical properties of actin over a wide range of oscillation frequencies and shear rates. Remarkably, both filamentous and nonfilamentous actin behaved as viscoelastic solids in both oscillatory and shear type experiments, providing that they were given ample time to equilibrate. Actin was purified by gel filtration from rabbit skeletal muscle and Acanthamoeba. Nonfilamentous actin in 2 different buffers had similar properties. In a low ionic strength buffer the absence of filaments was confirmed by electron microscopy, ultracentrifugation, and the fluorescence of pyrene-labeled actin. In 0.6 M KI, actin was monomeric by gel filtration. Filamentous actin had similar properties in 2 mM MgCl2 with either 50 mM KC1 or 500 mM KC1. Under all 4 of these conditions, actin required about 1000 min at 25 degrees C for the rheological properties to equilibrate. Under conditions where the oscillation of the rheometer did not affect the mechanical properties, all of the actin preparations had dynamic viscosities that were inverse functions of the frequency and dynamic elasticites that leveled off at low frequencies as expected for viscoelastic solids. For filamentous actin, the values of these parameters were about 2 times higher than for nonfilamentous actin. In shear experiments, both filamentous and nonfilamentous actin exhibited shear rate-dependent yield stresses. When filamentous and nonfilamentous actin structures were disrupted by transient shearing, the dynamic elasticity recovered to 90% in 30 min. Ovalbumin in the low ionic strength buffer also behaved as a viscoelastic material with elasticity and viscosity about 10 times lower than nonfilamentous actin, while cytochrome c behaved as a Newtonian fluid with a viscosity of 0.02 poise.
Highlights
From the Departmentof Cell Biology and Anatomy and the $Departmenotf Chemical Engineering, Johns Hopkins University, Baltimore, Maryland 21205
In a low ionic strength buffer the absence of filaments was confirmed by electron microscopy, ultracentrifugation, and the fluorescence of pyrenein termsof dynamic elasticity(G’) and dynamicviscosity (9’) as a function of frequency (6, 7)
Under all 4 of these conditions, actin required about biological materials, this parameter is a complex function of 1000 min at 25 “C for the rheological properties to shear rate, which is determined by the rheometer geometry equilibrate
Summary
From the Departmentof Cell Biology and Anatomy and the $Departmenotf Chemical Engineering, Johns Hopkins University, Baltimore, Maryland 21205. For. Under all 4 of these conditions, actin required about biological materials, this parameter is a complex function of 1000 min at 25 “C for the rheological properties to shear rate, which is determined by the rheometer geometry equilibrate. All of the actin preparations had dynamic viscosities Dynamic viscosity ( q ’ ) anddynamicelasticity (G’) are that were inverse functionsof the frequency and dy- parameters measured in oscillatory experiments (10-12). We have begun by characterizing solutionsof puritended toward infinity laotw shear rate These investigations suggested thatthefilaments formed weak, shear-sensitive networks that might or might not be cross-linked. They performed oscillatory experimentsandnoted afrequencydependent rigidity (0.15 dyne/cm* for 1 mg/ml) which they characterized over a limited range (0.017 to 0.17 Hz) with the parameter, dynamic elasticmodulus.
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