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  • Actin filament dynamics in the actomyosin VI complex is regulated allosterically by calcium-calmodulin light chain.

Actin filament dynamics in the actomyosin VI complex is regulated allosterically by calcium-calmodulin light chain.

Journal of molecular biology (2011-09-14)
Ewa Prochniewicz, Anaëlle Pierre, Brannon R McCullough, Harvey F Chin, Wenxiang Cao, Lauren P Saunders, David D Thomas, Enrique M De La Cruz
ABSTRACT

The contractile and enzymatic activities of myosin VI are regulated by calcium binding to associated calmodulin (CaM) light chains. We have used transient phosphorescence anisotropy to monitor the microsecond rotational dynamics of erythrosin-iodoacetamide-labeled actin with strongly bound myosin VI (MVI) and to evaluate the effect of MVI-bound CaM light chain on actin filament dynamics. MVI binding lowers the amplitude but accelerates actin filament microsecond dynamics in a Ca(2+)- and CaM-dependent manner, as indicated from an increase in the final anisotropy and a decrease in the correlation time of transient phosphorescence anisotropy decays. MVI with bound apo-CaM or Ca(2+)-CaM weakly affects actin filament microsecond dynamics, relative to other myosins (e.g., muscle myosin II and myosin Va). CaM dissociation from bound MVI damps filament rotational dynamics (i.e., increases the torsional rigidity), such that the perturbation is comparable to that induced by other characterized myosins. Analysis of individual actin filament shape fluctuations imaged by fluorescence microscopy reveals a correlated effect on filament bending mechanics. These data support a model in which Ca(2+)-dependent CaM binding to the IQ domain of MVI is linked to an allosteric reorganization of the actin binding site(s), which alters the structural dynamics and the mechanical rigidity of actin filaments. Such modulation of filament dynamics may contribute to the Ca(2)(+)- and CaM-dependent regulation of myosin VI motility and ATP utilization.

MATERIALS
Product Number
Brand
Product Description

Supelco
Erythrosin B, analytical standard
Sigma-Aldrich
Erythrosin extra bluish, for microscopy (Bact., Hist.), adsorption and fluorescent indicator
Sigma-Aldrich
Erythrosin B, certified by the Biological Stain Commission, Dye content ≥85 %
Sigma-Aldrich
Erythrosin extra bluish, certified by the Biological Stain Commission