CHARACTERIZATION OF APOCYTOCHROME-C BINDING TO HUMAN ERYTHROCYTES

The binding of I-125-labeled apocytochrome c to human erythrocytes was determined for free apocytochrome c concentrations at 10(-10)-10(-6) M. At about 2 x 10(-9) M, maximum cell association of apocytochrome c occurs at 50 mM NaCl and at 22 degrees C. Intact erythrocytes at 22 de grees C have three classes of apocytochrome c binding sites: one high- affinity noncooperative site(n(1), = 728 per cell, K-d1 = 1.5 x 10(-9) M) and two positively cooperative sites (n(2) = 3.7 x 10(4) per cell, K-d2 = 1.2 x 10(-7) M, alpha(2) = 2.0, and n(3) = 2.5 x 10(5) per cel l, K-d3 = 7.1 x 10(-7) M, alpha(3) = 12). Erythrocytes at 37 degrees C , and erythrocyte ghosts at 22 degrees C, also have three classes of a pocytochrome c binding sites, and most sites are positively cooperativ e. (C) 1994 Wiley-Liss, Inc.