LOW-K(M) AND HIGH-K(M) TRANSPORT OF DINITROPHENYL GLUTATHIONE IN INSIDE OUT VESICLES FROM HUMAN ERYTHROCYTES

Kinetic studies on the low- and high-K(m) transport systems for S-2,4-dinitrophenyl glutathione (DNP-SG) present in erythrocyte membranes were performed using inside-out plasma membrane vesicles. The high-affinity system showed a K(m) of 3.9-mu-M a V(max) of 6.3 nmol/mg protein per h, and the low-affinity system a K(m) of 1.6 mM and a V(max) of 131 nmol/mg protein per h. Both uptake components were inhibited by fluoride, vanadate, p-chloromercuribenzoate (pCMB) and bis(4-nitrophenyl)dithio-3,3'-dicarboxylate (DTNB). The low-K(m) uptake process was less sensitive to the inhibitory action of DTNB as compared to the high-K(m) process. N-Ethylmaleimide (1 mM) inhibited the high-K(m) process only. The high-affinity uptake of DNP-SG was competitively inhibited by GSSG (K(i) = 88-mu-M). Vice versa, DNP-SG inhibited competitively the low-K(m) component of GSSG uptake (K(i) = 3.3-mu-M). The high-K(m) DNP-SG uptake system was not inhibited by GSSG. The existence of a common high-affinity transporter for DNP-SG and GSSG in erythrocytes is suggested.