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Fig. 4. T-plastin or ABD1 stabilize actin filaments and protect them against cofilin-mediated depolymerization. G-Actin (4 µM, 25% pyrene labeled) was co-polymerized with various concentrations of T-plastin or ABD1. To induce depolymerization, T-plastin- or ABD1-decorated pyrene-labeled -F-actin was diluted under the critical concentration of the filament minus (–) end. Depolymerization of F-actin as a function of time was measured as a decrease in fluorescence because of the rapid depolymerization of actin filaments from their pointed ends. The fluorescence scale was adjusted so that the fluorescence of F-actin was 100 and that of G-actin was 0. (A) F-Actin (4 µM) co-polymerized with T-plastin (actin:T-plastin ratio of 1:1) was diluted to a concentration of 400 nM. Control (actin) was the depolymerization of F-actin in the absence of protein addition. (B) F-Actin decorated with ABD1 (actin:ABD1 ratios of 1:1, 1:2 or 1:4) was diluted to a concentration of 400 nM. Control (actin) was the depolymerization of F-actin in the absence of protein addition. (C) F-Actin decorated with T-plastin (4 µM) was diluted to a concentration of 400 nM in F buffer in the presence of 400 nM of GST-cofilin or its absence. Controls were the depolymerization of F-actin in the absence of T-plastin or GST-cofilin (Actin) or in the presence of 400 nM of GST-cofilin (Actin + cofilin). (D) F-Actin decorated with ABD1 (actin:ABD1 ratios of 1:2 or 1:4) was diluted to a concentration of 400 nM in the absence or presence of 400 nM of GST-cofilin. Controls as described in C.
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