IDENTIFICATION OF 2 SITES IN GELSOLIN WITH DIFFERENT SENSITIVITIES TOADENINE-NUCLEOTIDES

The affinity of monomeric actin for several actin-binding proteins, in cluding gelsolin, depends on adenine nucleotides. Gelsolin binds faste r and with higher affinity to ADP-actin than to ATP-actin. Here, we sh ow that the C-terminal actin-binding domain of gelsolin, which is requ ired for filament nucleating activity but not for filament severing ac tivity, 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 nucleo tide (ATP or ADP) tightly bound in the cleft of the actin monomer. Bin ding 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 forme d using different nucleotide concentrations differ in their filament-s evering activities as well as in their abilities to increase the fluor escence of 4-chloro-7-nitrobenzeno-2-oxa-1,3-diazole-labeled actin mon omers. These results suggest that, at physiologic concentrations of nu cleotides, both free and actin-bound ATP may affect the binding of act in to its accessory proteins and that gelsolin, actin, or the gelsolin -actin complex, contains a low-affinity nucleotide-binding site.