Synthesis of tritiated 4,4'-diisothiocyano-2,2'-stilbene disulfonic acid ([3H]DIDS) and its covalent reaction with sites related to anion transport in human red blood cells.
The potent and specific inhibitor of anion permeability, 4,4'-diisothicyanostilbene-2,2'-disulfonic acid (DIDS) was synthesized in tritiated form ([3H]DIDS) from tritiated 5-nitrotoluene-o-sulfonic acid. Its reactions with and effects on red blood cells were compared with those of a reduced form ([3H]H2DIDS), previously used as a tracer for DIDS. The rate of covalent reaction of [3H]DIDS was substantially faster than that of [3H]H2DIDS at all temperatures tested. With both agents, the rate of reaction was increased in alkaline media, although the response occurred at a lower pH with [3H]DIDS. On the other hand, the relationship of irreversible membrane binding to the degree of inhibition of sulfate fluxes was linear and virtually the same for both agents, with 100% inhibition associated with the binding of approximately 1.2 X 10(6) molecules per cell. About 90% of the binding for each probe was to a particular membrane protein, known as band 3, equivalent to about 1 mole of agent per mole of protein.
D000838:Anions; D001557:Benzenesulfonates / Q000138:chemical synthesis; D001665:Binding Sites; D001693:Biological Transport, Active; D002463:Cell Membrane Permeability; D004910:Erythrocyte Membrane / Q000378:metabolism*; D004912:Erythrocytes / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D013267:Stilbenes / Q000138:chemical synthesis*; D013431:Sulfates / Q000097:blood; D013696:Temperature; D013861:Thiocyanates / Q000138:chemical synthesis
D000838:Anions; D001557:Benzenesulfonates; D013267:Stilbenes; D013431:Sulfates; D013861:Thiocyanates