Amino acid residues complexed with eosin 5-isothiocyanate in band 3 protein of the human erythrocyte.
The amino-acid residues of band 3 protein taking part in the ionic interaction with an anion-transport inhibitor, eosin 5-isothiocyanate (EITC), were determined by pH titration. The plots of the absorbance of EITC-ghost system against pH reveal five equilibria, at pH 3.7, 6.4, 8.0, 11.0 and 13.1. Since the three equilibria, 3.7, 8.0 and 11.0, are representative of the EITC molecule, the others, 6.4 and 13.1, may be due to the interaction of EITC molecules with histidine and arginine residues, respectively. The same experiment using a reconstituted system of band 3-lipid vesicles gave results in good agreement with the EITC-ghost system. The intensity of the induced CD band at 530 nm of EITC molecules bound to ghosts was decreased by preincubation with arginine-specific reagents, phenylglyoxal and 1,2-cyclohexanedione, or histidine-specific reagents, diethylpyrocarbonate (DEPC) and p-diazobenzene sulfonate. The repression effects by these chemical modifiers were evaluated by measuring the concentrations which elicit 50% reduction. The histidine-specific reagents repressed the CD of EITC more effectively than the arginine-specific reagents. Furthermore, it was found that DEPC effectively inhibited the sulfate efflux from intact erythrocytes. These results suggest that the histidine residues participate in the anion-transport system of human red cells.
Chiba
T
T
Sato
Y
Y
Suzuki
Y
Y
D001457:Anion Exchange Protein 1, Erythrocyte / Q000378:metabolism; D001120:Arginine; D001665:Binding Sites; D001692:Biological Transport / Q000187:drug effects; D002942:Circular Dichroism; D004047:Diethyl Pyrocarbonate / Q000494:pharmacology; D004801:Eosine Yellowish-(YS) / Q000031:analogs & derivatives* / Q000494:pharmacology; D004910:Erythrocyte Membrane / Q000378:metabolism; D006639:Histidine; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007473:Ion Channels / Q000187:drug effects; D008081:Liposomes; D011487:Protein Conformation; D013057:Spectrum Analysis
D001457:Anion Exchange Protein 1, Erythrocyte; D007473:Ion Channels; D008081:Liposomes; D006639:Histidine; C012957:eosine-5-isothiocyanate; D001120:Arginine; D004047:Diethyl Pyrocarbonate; D004801:Eosine Yellowish-(YS)