Two steps in ATP-dependent shape change of human erythrocyte ghosts

Most human erythrocyte membranes (ghosts) prepared with 10 mM Tris-Cl buffer were spherocytic and changed shape through crenated to discoidal in the presence of 2 mM Mg-ATP at 37 degrees C during 30 min under hypotonic conditions. These discocytic ghosts reverted to spherical form after being washed with 1 mM MgCl2, although their membranes were phosphorylated and they were converted to discocytes again, immediately on addition of 2 mM Mg-ATP, even at 6 degrees C. There seem to be 2 steps in the shape change of the ghosts; the first step proceeds gradually during incubation at 37 degrees C for 30 min in the presence of a physiological concentration of Mg-ATP and the second step occurs rapidly after addition of Mg-ATP even at 6 degrees C. This suggests that not only membrane phosphorylation but also specific ATP-binding (or hydrolysis) is necessary for erythrocyte shape change.