Studies on inactivation of anion transport in human red blood cell membrane by reversibly and irreversibly acting arginine-specific reagents.
A chromophoric derivative of phenylglyoxal, 4-hydroxy-3-nitrophenylglyoxal (HNPG), known to be highly selective for modification of arginine residues in aqueous solution is found to be a potent inhibitor of anion transport across the red cell membrane. In contrast to the action of all other arginine-specific reagents used under the experimental conditions in this laboratory, the action of HNPG on sulfate transport is completely reversible. Hence, a kinetic analysis of its inhibitory effect on SO4(2-) self-exchange could be performed. The effect of increasing chloride concentration on the inhibitory potency of HNPG is consistent with the concept that Cl- and HNPG compete for the same site on the anion transporter. The IC50 value for the inhibition of SO4(2-) exchange with HNPG is about 0.13 mM at pH 8.0 and 0.36 mM at pH 7.4, and the Hill coefficient for the interaction between the transporter and the inhibitor is near one at both pH's. HNPG is able to protect the transport system against inhibition with the (under our experimental conditions) irreversibly acting arginine specific reagent, phenylglyoxal. Partial inactivation of the transport system with phenylglyoxal lowers the maximal rates of SO4(2-) and chloride exchange but does not modify the apparent KS for the substrate anions. Reversibly acting anion transport inhibitors known to interact with the DIDS binding site like salicylate, tetrathionate, APMB, DNDS, and flufenamate are able to protect the transport system against phenylglyoxalation. Other inhibitors like phloretin and phlorizin have no effect.
Julien
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T
Max-Planck-Institut für Biophysik, Frankfurt, West Germany.
Zaki
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D000447:Aldehydes / Q000494:pharmacology*; D000838:Anions / Q000378:metabolism*; D001665:Binding Sites; D001692:Biological Transport / Q000187:drug effects; D002712:Chlorides / Q000378:metabolism; D004910:Erythrocyte Membrane / Q000187:drug effects* / Q000378:metabolism; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007474:Ion Exchange / Q000187:drug effects; D007700:Kinetics; D010658:Phenylglyoxal / Q000031:analogs & derivatives / Q000378:metabolism / Q000494:pharmacology*; D013431:Sulfates / Q000378:metabolism
D000447:Aldehydes; D000838:Anions; D002712:Chlorides; D013431:Sulfates; C027540:4-hydroxy-3-nitrophenylglyoxal; D010658:Phenylglyoxal