Impaired nicotinamide adenine dinucleotide synthesis in pyruvate kinase- deficient human erythrocytes: a mechanism for decreased total NAD content and a possible secondary cause of hemolysis

Erythrocytes from individuals with pyruvate kinase (PK) deficiency have approximately half the total (oxidized and reduced) nicotinamide adenine dinucleotide (NAD) of normal erythrocytes. In order to elucidate the mechanism(s) for the decrease in total NAD, we examined NAD synthesis in intact erythrocytes. It is demonstrated that NAD synthesis is impaired in PK-deficient erythrocytes to a degree that is dependent on the PK activity and adenosine 5'-triphosphate (ATP) concentration of these cells. After incubation in the presence of fluoride, which simulates the characteristics of PK deficiency by inhibiting enolase, normal erythrocytes had impaired NAD synthesis and decreased ATP concentrations. Fluoride did not inhibit NAD synthesis in a hemolysate system that is not dependent on glycolysis for ATP generation. These data suggest that fluoride does not inhibit the enzymes of NAD synthesis and that impairment of NAD synthesis by fluoride is mediated by decreased ATP formation. Thus, it is concluded that impaired NAD synthesis in PK-deficient erythrocytes is caused by decreased ATP formation due to the PK deficiency. Since the rate of glycolysis is limited by the availability of NAD+, it is suggested that impaired NAD synthesis causes further ATP depletion and thereby may enhance hemolysis in PK-deficient erythrocytes.