ROLE OF PROTEIN -SH GROUPS IN REDOX HOMEOSTASIS - THE ERYTHROCYTE AS A MODEL SYSTEM

The reactivities of the sulfhydryl groups of rat, turkey, human, and c alf hemoglobin were studied together with the enzyme activities of glu tathione peroxidase, glutathione reductase, glucose-6-phosphate dehydr ogenase, and glutaredoxin in lysed erythrocytes to evaluate their role s in regulating redox homeostasis. The results of -SH reactivity showe d rate constants spanning four orders of magnitude (k(2), calf, 6.67 M -1 s(-1); rat -SH fast reacting, 2.8 x 10(4) M-1 s(-1)). Enzyme activi ties of glucose-6-phosphate dehydrogenase ranged from 0.402 U/ml (calf ) to 0.900 U/ml (rat), glutathione reductase from 0.162 U/ml (rat) to 0.381 U/ml (human), glutaredoxin from 0.778 U/ml (rat) to 2.28 U/ml (t urkey), and glutathione peroxidase from 2.07 U/ml (human) to 27.3 U/ml (rat). Blood samples of the four species were also treated with 0.5-1 .5 mM tert-butyl hydroperoxide (t-BOOH) or diamide, and levels of glut athione-derived species [GSH, GSSG;, and glutathione-protein mixed dis ulfides (GS-SP)I were determined within 120 min and related to the cor responding protein -SH group (PSH) reactivities and enzyme repertoires . In all cases t-BOOH rapidly transformed GSH into GSSG; by the action of glutathione peroxidase; GSSG was in turn transformed into GS-SP, a ccording to the reaction GSSG + PSH --> GS-SP + GSH, or reduced back t o GSH by glutathione reductase. The GSSG reduction was more efficient in rat and human blood, due to the contribution of the fast-reacting - SH of hemoglobin, in the rat, and to the efficiency of the enzyme repe rtoire of human blood. Calf blood showed a relatively low capacity to restore normal values after oxidative stress, due to its low PSH react ivity and the weak contribution of its enzymes. Diamide treatment, whi ch is known to react nonenzymatically with thiols, gave increased GS-S P levels in rat and turkey, but not in human and calf blood, as expect ed from the different corresponding PSH reactivities. Species with rel atively high PSH reactivity and glucose 6-phosphate dehydrogenase acti vity, such as the rat, therefore had a higher antioxidant capacity tha n species (calf) in which these parameters were relatively low. (C) 19 98 Academic Press.