Purification and properties of Penicillium glucose 6-phosphate dehydrogenase.
1. Glucose 6-phosphate dehydrogenase was isolated and partially purified from a thermophilic fungus, Penicillium duponti, and a mesophilic fungus, Penicillium notatum. 2. The molecular weight of the P. duponti enzyme was found to be 120000+/-10000 by gelfiltration and sucrose-density-gradient-centrifugation techniques. No NADP(+)- or glucose 6-phosphate-induced change in molecular weight could be demonstrated. 3. Glucose 6-phosphate dehydrogenase from the thermophilic fungus was more heat-stable than that from the mesophile. Glucose 6-phosphate, but not NADP(+), protected the enzyme from both the thermophile and the mesophile from thermal inactivation. 4. The K(m) values determined for glucose 6-phosphate dehydrogenase from the thermophile P. duponti were 4.3x10(-5)m-NADP(+) and 1.6x10(-4)m-glucose 6-phosphate; for the enzyme from the mesophile P. notatum the values were 6.2x10(-5)m-NADP(+) and 2.5x10(-4)m-glucose 6-phosphate. 5. Inhibition by NADPH was competitive with respect to both NADP(+) and glucose 6-phosphate for both the P. duponti and P. notatum enzymes. The inhibition pattern indicated a rapid-equilibrium random mechanism, which may or may not involve a dead-end enzyme-NADP(+)-6-phosphogluconolactone complex; however, a compulsory-order mechanism that is consistent with all the results is proposed. 6. The activation energies for the P. duponti and P. notatum glucose 6-phosphate dehydrogenases were 40.2 and 41.4kJ.mol(-1) (9.6 and 9.9kcal.mol(-1)) respectively. 7. Palmitoyl-CoA inhibited P. duponti glucose 6-phosphate dehydrogenase and gave an inhibition constant of 5x10(-6)m. 8. Penicillium glucose 6-phosphate dehydrogenase had a high degree of substrate and coenzyme specificity.
D002499:Centrifugation, Density Gradient; D002729:Chloromercuribenzoates; D002850:Chromatography, Gel; D004591:Electrophoresis, Polyacrylamide Gel; D005954:Glucosephosphate Dehydrogenase / Q000037:antagonists & inhibitors / Q000302:isolation & purification*; D005958:Glucosephosphates; D007700:Kinetics; D008970:Molecular Weight; D009249:NADP; D010407:Penicillium / Q000201:enzymology*; D013395:Sucrose; D013696:Temperature
D002729:Chloromercuribenzoates; D005958:Glucosephosphates; D009249:NADP; D013395:Sucrose; D005954:Glucosephosphate Dehydrogenase