Dinitrophenyl glutathione efflux from human erythrocytes is primary active ATP-dependent transport

E. F. LaBelle, S. V. Singh, S. K. Srivastava, Y. C. Awasthi

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Abstract

Dinitrophenyl S-glutathione is accumulated by inside-out vesicles made from human erythrocytes in a process totally dependent on ATP and Mg2+. The vesicles were shown to accumulate dinitrophenyl S-glutathione against a concentration gradient. The vesicles were able to concentrate this glutathione derivative even in the absence of membrane potential. This indicated that the ATP-dependent uptake of dinitrophenyl S-glutathione by inside-out vesicles represented an active transport process. Neither extravesicular EGTA nor intravesicular ouabain inhibited the transport process, indicating that neither the Ca2+-ATPase nor the Na+,K+-ATPase were involved. These results indicated that dinitrophenyl S-glutathione uptake by inside-out vesicles probably represented primary active transport. The uptake of dinitrophenyl S-glutathione was a linear function of time (up to 5 h) and vesicle protein. The rate of uptake was optimal between pH 7.0 and 8.0 and at 37°C. The K(m) values determined for dinitrophenyl S-glutathione and ATP were 0.29 mM and 1 mM, respectively. The transport process was completely inhibited by vanadate and by p-hydroxymercuribenzene sulphonate and inhibited to a lesser extent by N-ethylmaleimide. GTP could efficiently substitute for ATP as an energy source for the transport process, but CTP and UTP were comparatively much less effective.

Original languageEnglish (US)
Pages (from-to)443-449
Number of pages7
JournalBiochemical Journal
Volume238
Issue number2
DOIs
StatePublished - Jan 1 1986

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ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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