Kinetics of DIDS inhibition of swelling-activated K-Cl cotransport in low K sheep erythrocytes.
The inhibitory effect of various stilbene disulfonates was examined on the swelling-activated Cl-dependent K transport (K-Cl cotransport) in low K sheep erythrocytes. Both diisothiocyanatostilbenes H2DIDS and DIDS were found to be potent inhibitors. The DIDS concentration yielding 50% inhibition (IC50) of KCl cotransport was 60 microM in the absence of external K and 3 microM at physiological K concentration. Other stilbene derivatives, such as SITS (4-acetamido-4' isothiocyanatostilbene-2,2'-disulfonic acid), were only effective in the presence of external K, whereas DNDS (4,4'-dinitrostilbene-2,2'-disulfonic acid) and ISA (4-sulfophenyl isothiocyanate) had only slight effects at a concentration of 1 mM. The augmenting effect of external K is due to a second K site, distinguishable from the K transport site by its much higher affinity. No inhibition occurred in the absence of external Cl, whether or not external Rb(K) was present. Additionally, DIDS inhibited K-Cl cotransport activated by thiol alkylation with N-ethylmaleimide (NEM) as well as by Mg depletion in the presence of A23187 and a chelator. We conclude that allosteric sites affect the stilbene binding. When these sites are saturated, changes in external K or Cl concentration do not affect the affinity for DIDS (noncompetitive inhibition).
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Department of Physiology and Biophysics, School of Medicine, Wright State University, Dayton, Ohio 45401-0927.
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D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid; D012856:4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic Acid / Q000031:analogs & derivatives* / Q000494:pharmacology; D000818:Animals; D001692:Biological Transport / Q000187:drug effects / Q000502:physiology; D000001:Calcimycin / Q000494:pharmacology; D002352:Carrier Proteins / Q000037:antagonists & inhibitors* / Q000502:physiology; D002712:Chlorides / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004907:Erythrocyte Deformability / Q000187:drug effects*; D004912:Erythrocytes / Q000737:chemistry / Q000378:metabolism* / Q000648:ultrastructure; D005033:Ethylmaleimide / Q000494:pharmacology; D011188:Potassium / Q000032:analysis / Q000378:metabolism / Q000493:pharmacokinetics*; D012756:Sheep; D013267:Stilbenes / Q000494:pharmacology; D027981:Symporters; D000096922:K Cl- Cotransporters
D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid; D012856:4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic Acid; D000001:Calcimycin; D002352:Carrier Proteins; D002712:Chlorides; D005033:Ethylmaleimide; D011188:Potassium; D013267:Stilbenes; D027981:Symporters; D000096922:K Cl- Cotransporters; C022306:4,4'-dinitro-2,2'-stilbenedisulfonic acid