Tissue Expression and Actin Binding of a Novel N-Terminal Utrophin Isoform
Figure 6
Binding of utrophin and dystrophin fragments to actin by cosedimentation. Assays were carried out in 2 mM MgCl2 and 0.5 mM CaCl2 (O) or 6 mM MgCl2 and 5 mM EGTA (●). Saturation curves for binding of UT11 (a), UT12 (c), and DYS12 (e) at the left side with the corresponding Scatchard plots at the right (b, d, f). The binding curves resulted from 7 separate experiments each. All data points (●) are incorporated in the Scatchard plots except for DYS12 in the presence of EGTA. The concave Scatchard curves (b, d) indicate the presence of a higher and a lower affinity due to either two classes of binding sites or, alternatively, negative cooperativity. The convex curve (f) suggests positive cooperativity of DYS12 binding. Binding of DYS12 to actin was also measured with increasing concentrations of EGTA (g) in the presence of 1 mM CaCl2 in excess over the EGTA. Numbers at data points refer to the number of experiments. Error bars indicate SEM with for 0 and 0.5 versus 1.0 and 5.0 mM EGTA. The small differences between 0 versus 0.5 and 1.0 versus 5.0 mM EGTA were not significant. For further explanation, see the text.
