DIVALENT-CATION BINDING TO ERYTHROCYTE SPECTRIN

Erythrocyte spectrin dimers and separated alpha- and beta-spectrin cha ins bound (Ca2+)-Ca-45 after electrophoresis on native or sodium dodec yl sulfate-polyacrylamide gels, blotting, and (Ca2+)-Ca-45 overlay. Fl ow dialysis and equilibrium dialysis revealed two binding components: high-affinity, Ca2+-specific sites with k(d) = 4 x 10(-7) M and n = 10 0 +/- 20 per dimer and a low-affinity (millimolar) divalent cation com ponent. Whereas brain spectrin had only four high-affinity sites [Wall is, C. J., Wenegieme, E. F., & Babitch, J. A. (1992) J. Biol. Chem. 26 7, 4333-4337], erythrocyte spectrin had 25-fold more sites per dimer. In addition to possibly modifying spectrin interactions with calcium-d ependent protease and actin, as suggested by previous work on the inte raction of Ca2+ with brain spectrin, the approximately two high-affini ty sites per repeating segment of erythrocyte spectrin appear to stabi lize a folded conformation of repeat structures indicated by an entrop y increase upon binding. These data support the hypothesis that divale nt cation binding to erythrocyte spectrin has become specialized to st abilize the membrane skeletal network and the cell, making them flexib le but resistant to shear under the stressful conditions of blood circ ulation.