Ornithine decarboxylase in mouse kidney. Purification, characterization, and radioimmunological determination of the enzyme protein.
Ornithine decarboxylase was purified from kidneys of androgen-treated mice to a greater than 95% purity. Two-dimensional gel electrophoresis revealed charge heterogeneity in the enzyme (pI 4.7-4.9) which was observed both by protein staining and by covalent labeling with [3H]alpha-difluoromethylornithine. Antibodies raised in rabbits inhibited the activity of the enzyme, formed a single rocket in crossed immunoelectrophoresis, and bound [3H]alpha-difluoromethylornithine-labeled enzyme as well as [125I]iodoornithine decarboxylase. A sensitive radioimmunoassay for the enzyme was established; the minimal detectable enzyme concentration was 0.1 ng/assay tube that corresponded to about 0.1-0.2 ng/mg of cytosol protein. The antiserum cross-reacted with enzymes from different tissues from mouse, rat, hamster, and human. Immunoreactive ornithine decarboxylase concentration in renal cytosol of male mice was 13-fold higher than that in the females (36.2 +/- 2.7 versus 2.8 +/- 0.2 ng/mg of cytosol protein; mean +/- S.E.); treatment with testosterone implants for 5-7 days increased the concentration to 522 +/- 66 ng/mg of protein. After administration of a single dose of testosterone (10 mg) to female mice, an increased immunoreactive ornithine decarboxylase concentration was detected as soon as 2 h, and rose sharply between 8 and 24 h after steroid administration. These changes were similar to those seen by assays of the catalytically active enzyme. The half-life of immunoreactive ornithine decarboxylase in mouse kidney, as measured after inhibition of protein synthesis in vivo by cycloheximide administration, was 16 min in nontreated and 140 min in androgen-treated male animals, while the corresponding values for the catalytically active enzyme were 9 and 90 min.
D000818:Animals; D000936:Antigen-Antibody Complex; D002262:Carboxy-Lyases / Q000302:isolation & purification*; D003600:Cytosol / Q000201:enzymology; D004343:Drug Implants; D005260:Female; D007106:Immune Sera; D007668:Kidney / Q000187:drug effects / Q000201:enzymology*; D007700:Kinetics; D051379:Mice; D009955:Ornithine Decarboxylase / Q000302:isolation & purification* / Q000378:metabolism; D013739:Testosterone / Q000494:pharmacology*
D000936:Antigen-Antibody Complex; D004343:Drug Implants; D007106:Immune Sera; D013739:Testosterone; D002262:Carboxy-Lyases; D009955:Ornithine Decarboxylase