The kinetic properties of placental glucose-6-phosphate dehydrogenase were studied, since this enzyme is expected to be an important component of the placental protection system. In this capacity it is also very important for the health of the fetus. The placental enzyme obeyed "Rapid Equilibrium Ordered Bi Bi'' sequential kinetics with K-m:, values of 40 +/- 8 muM for glucose-6-phosphate and 20 +/- 10 muM for NADP. Glucose-6-phosphate, 2-deoxyglucose-6-phosphate and galactose-6-phosphate were used with catalytic efficiencies (k(cat)/K-m) of 7.4 x 10(6), 4.89 x 10(4) and 1.57 x 10(4) M-1.s(-1). respectively, The K(m)app values for galactose-6-phosphate and for 2-deoxyglucose-6-phosphate were 10 +/- 2 and 0.87 +/- 0.06 mM. With galactose-6-phosphate as substrate, the same K-m, value for NADP as glucose-6-phosphate was obtained and it was independent of galactose-6-phosphate concentration. On the other hand, when 2-deoxyglucose-6-phosphate used as substrate, the ii:, for NADP decreased from 30 +/- 6 to 10 +/- 2 muM as the substrate concentration was increased from 0.3 to 1.5 mM. Deamino-NADP, but not NAD, was a coenzyme for placental glucose-6-phosphate dehydrogenase. The catalytic efficiencies of NADP and deamino-NADP (glucose-6-phosphate as substrate) were 1.48 x 10(7) and 4.80 x 10(6) M(-1)s(-1), respectively. With both coenzymes, a hyperbolic saturation and an inhibition above 300 muM coenzyme concentration, was observed. Human placental glucose-6-phosphate dehydrogenase was inhibited competitively by 2,3-diphosphoglycerate (K-i = 15 +/- 3 mM) and NADPH (K-i = 17.1 +/- 3.2 muM). The small dissociation constant for the G6PD:NADPH complex pointed to tight enzyme:NADPH binding and the important role of NADPH in the regulation of the pentose phosphate pathway. (C) 2001 Elsevier Science Ltd. All lights reserved.