Mechanisms of decreased erythrocyte deformability and survival in glucose 6-phosphate dehydrogenase mutants.

T. P. Flynn, G. J. Johnson, D. W. Allen

Research output: Contribution to journalArticle

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Abstract

We have studied the nature of the oxidative lesion of the erythrocyte membrane in glucose-6-phosphate dehydrogenase (G6PD) mutants with chronic hemolysis, comparing these membranes with those from normal red cells (RBC) subjected to oxidative stress in vitro. Disulfide-linked polypeptide aggregates are found in membranes from fresh RBC of these G6PD mutants and from aerobically incubated normal erythrocytes. As further evidence of oxidative damage, increased disulfide bonds were found in the RBC membranes from both the mutants and incubated normal RBC. The intermolecular bonds which cross-link membrane polypeptides to form the observed aggregates, however, only accounted for a fraction of the membrane disulfide bonds present. Thus, most of the disulfide bonds in the G6PD mutants were intramolecular. These intramolecular disulfide bonds were widely distributed on the membrane polypeptides, but were found to be concentrated on cytoskeletal anchoring proteins, bands 2.1-2.3, using [14C] iodoacetamide labelling of the sulfhydryls involved in disulfide bonds. The intermolecular bonds, on the other hand, were concentrated in spectrin. When G6PD mutant membranes were examined on sucrose density gradients, a subpopulation of dense membranes was observed which resembled the membranes of oxidatively stressed normal RBC both in increased density and in increased binding of nonhemoglobin cytoplasmic protein. To study the relationship between sulfhydryl oxidation, membrane density and RBC viscosity the sulfhydryl oxidant diamide (diazine dicarboxylic acid bis-[dimethylamide]) was used. Diamide treated erythrocytes, like the G6PD mutants, had decreased GSH, increased polypeptide aggregates, increased viscosity, but no change in ATP. We conclude that in G6PD mutants with chronic hemolysis oxidative damage includes aggregate formation due to intermolecular disulfide bonds, and intramolecular disulfide bond formation associated with increased binding of non-hemoglobin cytoplasmic proteins to the membrane. The relative importance of intermolecular and intramolecular disulfide bond formation and the mechanism whereby these changes may produce decreased RBC deformability and survival remain to be determined.

Original languageEnglish (US)
Pages (from-to)231-249
Number of pages19
JournalProgress in clinical and biological research
Volume56
StatePublished - 1981

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