Divalent cation binding to erythrocyte spectrin.
Erythrocyte spectrin dimers and separated alpha- and beta-spectrin chains bound 45Ca2+ after electrophoresis on native or sodium dodecyl sulfate-polyacrylamide gels, blotting, and 45Ca2+ overlay. Flow dialysis and equilibrium dialysis revealed two binding components: high-affinity, Ca(2+)-specific sites with kd = 4 x 10(-7) M and n = 100 +/- 20 per dimer and a low-affinity (millimolar) divalent cation component. Whereas brain spectrin had only four high-affinity sites [Wallis, C. J., Wenegieme, E. F., & Babitch, J. A. (1992) J. Biol. Chem. 267, 4333-4337], erythrocyte spectrin had 25-fold more sites per dimer. In addition to possibly modifying spectrin interactions with calcium-dependent protease and actin, as suggested by previous work on the interaction of Ca2+ with brain spectrin, the approximately two high-affinity sites per repeating segment of erythrocyte spectrin appear to stabilize a folded conformation of repeat structures indicated by an entropy increase upon binding. These data support the hypothesis that divalent cation binding to erythrocyte spectrin has become specialized to stabilize the membrane skeletal network and the cell, making them flexible but resistant to shear under the stressful conditions of blood circulation.
Department of Chemistry, Texas Christian University, Forth Worth 76129.
C J
CJ
Wallis
J A
JA
Babitch
E F
EF
Wenegieme
D000818:Animals; D001665:Binding Sites; D002118:Calcium / Q000097:blood*; D002132:Calcium Radioisotopes; D002413:Cations, Divalent; D002942:Circular Dichroism; D003956:Dialysis; D004591:Electrophoresis, Polyacrylamide Gel; D004912:Erythrocytes / Q000737:chemistry*; D006736:Horses; D006801:Humans; D046911:Macromolecular Substances; D015636:Magnesium Chloride / Q000494:pharmacology; D011189:Potassium Chloride / Q000494:pharmacology; D013049:Spectrin / Q000737:chemistry / Q000378:metabolism*
D002132:Calcium Radioisotopes; D002413:Cations, Divalent; D046911:Macromolecular Substances; D015636:Magnesium Chloride; D013049:Spectrin; D011189:Potassium Chloride; D002118:Calcium