Kinetics of DIDS inhibition of swelling-activated K-Cl cotransport in low K sheep erythrocytes

• Published in: The Journal of membrane biology 1992-02, Vol.126 (1), p.89-96
• Main Authors: DELPIRE, E, LAUF, P. K
• Format: Article
• Language: English
• Subjects: 4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid; 4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic Acid - analogs & derivatives; 4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic Acid - pharmacology; Animals; Biological and medical sciences; Biological Transport - drug effects; Biological Transport - physiology; Calcimycin - pharmacology; Carrier Proteins - antagonists & inhibitors; Carrier Proteins - physiology; Cell physiology; chloride; Chlorides - pharmacology; Dose-Response Relationship, Drug; Erythrocyte Deformability - drug effects; erythrocytes; Erythrocytes - chemistry; Erythrocytes - metabolism; Erythrocytes - ultrastructure; Ethylmaleimide - pharmacology; Fundamental and applied biological sciences. Psychology; inhibition; K Cl- Cotransporters; kinetics; Membrane and intracellular transports; Molecular and cellular biology; potassium; Potassium - analysis; Potassium - metabolism; Potassium - pharmacokinetics; Sheep; Stilbenes - pharmacology; swelling; Symporters; transport
• ISSN: 0022-2631; 1432-1424

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).