Identification of two sites in gelsolin with different sensitivities to adenine nucleotides.
The affinity of monomeric actin for several actin-binding proteins, including gelsolin, depends on adenine nucleotides. Gelsolin binds faster and with higher affinity to ADP-actin than to ATP-actin. Here, we show that the C-terminal actin-binding domain of gelsolin, which is required for filament nucleating activity but not for filament severing activity, contains the site that distinguishes between ATP-actin and ADP-actin monomers. In contrast, actin binding to the N-terminal half of gelsolin depends on solution ATP concentrations, but not on the nucleotide (ATP or ADP) tightly bound in the cleft of the actin monomer. Binding is stronger in the absence of free nucleotide or in the presence of 0.5 mM ADP than in solutions containing 0.5 mM ATP. Complexes formed using different nucleotide concentrations differ in their filament-severing activities as well as in their abilities to increase the fluorescence of 4-chloro-7-nitrobenzeno-2-oxa-1,3-diazole-labeled actin monomers. These results suggest that, at physiologic concentrations of nucleotides, both free and actin-bound ATP may affect the binding of actin to its accessory proteins and that gelsolin, actin, or the gelsolin-actin complex, contains a low-affinity nucleotide-binding site.
Department of Medicine, Brigham and Women's Hospital, Boston, MA, USA.
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Laham
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Way
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Yin
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Janmey
D000199:Actins / Q000378:metabolism; D000244:Adenosine Diphosphate / Q000378:metabolism*; D000255:Adenosine Triphosphate / Q000378:metabolism*; D001665:Binding Sites; D018260:Gelsolin / Q000378:metabolism*; D006801:Humans
D000199:Actins; D018260:Gelsolin; D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate