Transduction of reducing power across the plasma membrane by reduced glutathione. A 1H-NMR spin-echo study of intact human erythrocytes

M. R. Ciriolo, M. Paci, M. Sette, A. De Martino, A. Bozzi, G. Rotilio

Research output: Contribution to journalArticlepeer-review

Abstract

The NMR signal of reduced glutathione (GSH) was monitored in intact human erythrocytes by the 1H spin-echo Carr-Purcell-Meiboom-Gill pulse sequence. Addition of GSH, which was unable to cross the erythrocyte membrane, produced an approximate two-fold increase of the GSH signal in glucose-depleted cells. Addition of oxidised glutathione (GSSG), did not affect the signal, and addition of GSH to hemolysates gave a much smaller increase. Reduction of internal GSSG by NADPH-dependent enzymes was excluded by experiments with glucose-supplied or glucose-6-phosphate dehydrogenase deficient cells. Involvement of external thiol groups of the erythrocyte membrane was shown by the lack of effect in cells treated with an impermeable thiol-blocking compound. Involvement of spectrin was indicated by the proportional loss of the effect in erythrocytes with variable genetic deficiency of spectrin. Protein-glutathione mixed disulfides appeared to be the source of the NMR response since an increase of their content, by diamide treatment or aging procedures, produced a higher GSH signal, while their reduction by permeable reductants gave the opposite effect. It is concluded that GSH can transduce its reducing power by a thiol/disulfide exchange mechanism that sequentially involves sulfur-rich proteins spanning across the erythrocyte membrane.

Original languageEnglish
Pages (from-to)711-718
Number of pages8
JournalEuropean Journal of Biochemistry
Volume215
Issue number3
Publication statusPublished - 1993

ASJC Scopus subject areas

  • Biochemistry

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