Figure 2.
High-affinity binding of dendritic BC1 RNA to hnRNP A2 is modulated by Mg2+ and Ca2+ ions. Binding affinities were established by quantitative EMSA using native PAGE. WT BC1 RNA (A) and WC mutant BC1 RNA (B) were titrated with increasing concentrations of hnRNP A2 in the presence of 10 mM MgCl2. Although a Kd of 8.2 nM was obtained for WT BC1 RNA, no equilibrium constant could be established for WC mutant BC1 RNA, as no binding was observed even at maximal hnRNP A2 concentrations. (C) Binding affinity of BC1 RNA to hnRNP A2 is dependent on the concentration of Mg2+ ions. The following equilibrium dissociation constants were obtained: 25 mM MgCl2, Kd = 3.5 nM; 10 mM MgCl2, Kd = 8.2 nM; 5 mM MgCl2, Kd = 520 nM; 2.5 mM MgCl2, Kd = 980 nM. (D) Binding of BC1 RNA to hnRNP A2 is modulated by Ca2+ ion concentrations. Dissociation constants: 10 mM CaCl2, Kd = 0.6 nM; 5 mM CaCl2, Kd = 1.0 nM; 1 mM CaCl2, Kd = 210 nM; 0.5 mM CaCl2, Kd = 250 nM; 0.1 mM CaCl2, Kd = 380 nM.