PURIFICATION, CHARACTERIZATION AND KINETIC MECHANISM OF GLUCOSE-6-PHOSPHATE-DEHYDROGENASE FROM MOUSE-LIVER

1. Glucose-6-phosphate dehydrogenase (G6PDH EC 1.1.1.49) from mouse li ver has been purified 1100-fold by extraction, ion-exchange chromatogr aphy on DE-52, absorption chromatography on Bio-Gel HTP and gel filtra tion through sepharose 6 HR 10/30. The purified enzyme showed a single band in silver stained SDS-PAGE. 2. The native and subunit molecular weight were 117 and 31 kDa respectively. 3. The kinetic studies and th e patterns obtained from the inhibition by-products suggest that the e nzyme follows an ordered sequential kinetic mechanism. 4. The reduced K-m values for the substrates favour the operativity of the enzyme. Th e ''fine control'' of the enzymatic activity was exerted by the NADPH, whose Ki is several fold lower than the in vivo concentration.