High affinity L-triiodothyronine binding to right-side-out and inside-out vesicles from rat and human erythrocyte...

3,5,3'-Triiodo-L-thyronine (L-T3)-binding sites from rat and human red cells were characterized as to their distribution between the two surfaces of the membrane. Analysis of L-T3 binding to sealed right-side-out and inside-out vesicles from erythrocyte membrane revealed that high affinity L-T3-binding sites are located on the external side in rat erythrocytes and on the internal side in human red cells. These results were further confirmed by preincubation of intact red cells with p-chloromercuribenzoate, a slowly permeant reagent that interacts reversibly with SH groups of proteins. Following this treatment only the SH groups of L-T3 sites from rat erythrocytes were found to be blocked. Scatchard analysis of the binding data for rat right-side-out and human inside-out vesicles showed high affinity sites with Kd values of 0.2 x 10(-10) and 2 x 10(-10) M, respectively. The results suggest that the orientation of L-T3-binding sites in the erythrocyte membrane is species-dependent.