title: Purification and characterization of mouse glucose 6-phosphate dehydrogenase.
abstract: Glucose-6-phosphate dehydrogenase was purified to homogeneity from testes and kidneys of the inbred strain of mice (DBA/2J) by a simple two-step affinity column procedure. This involved the sequential application of 8-(6-aminohexyl)-amino-AMP- and -2', 5'-ADP-Sepharose columns and biospecific elution with NADP+ in both steps. The molecular and biochemical properties of the purified enzyme were studied in detail. These include the molecular weight determination, amino acid composition, steady-state kinetics, inactivation by high temperature, urea and iodoacetate, and immunology. The purified enzyme from mouse kidneys or testes was shown to be a tetramer with a molecular weight of 220,000. The enzyme is highly specific for glucose-6-phosphate, exhibits almost no activity with NAD+ as a coenzyme and is little inhibited by AMP or ATP. Michaelis constants for glucose-6-phosphate and NADP+ were determined to be 50 microM and 10 microM respectively. NADPH is a competitive inhibitor of NADP+ and has a Ki of 18 microM. Rabbit antisera against glucose-6-phosphate dehydrogenase were raised. The antisera also cross-react with the same enzyme from human and guinea pig.
mesh_terms: D000596:Amino Acids / Q000032:analysis; D000818:Animals; D005954:Glucosephosphate Dehydrogenase / Q000302:isolation & purification* / Q000378:metabolism; D007118:Immunoassay; D007668:Kidney / Q000201:enzymology*; D007700:Kinetics; D046911:Macromolecular Substances; D008297:Male; D051379:Mice; D008811:Mice, Inbred DBA; D008970:Molecular Weight; D013737:Testis / Q000201:enzymology*
chemical_list: D000596:Amino Acids; D046911:Macromolecular Substances; D005954:Glucosephosphate Dehydrogenase