Carbonic anhydrase isoenzymes in the erythrocytes and uterus of the rabbit.
1. Two forms of the zinc-containing enzyme carbonic anhydrase (EC 4.2.1.1) were isolated from rabbit erythrocytes and two forms from rabbit uterine tissue (endometrium) in the progestational stage of pregnancy (days 6-8 of gestation). Separation of the isoenzymes was achieved by ion-exchange chromatography, preparative polyacrylamide-gel electrophoresis and isoelectric focusing. A comparison was made of the general properties and kinetic behaviour of the purified isoenzymes. 2. Although indistinguishable in terms of molecular weight and zinc content the isoenzymes were very different as catalysts of the hydration of carbon dioxide. The two erythrocyte isoenzymes, found in almost equal amounts, differed more than 100-fold in specific activity. Of the two isoenzymes prepared from either endometrial or entire uterine homogenates one was kinetically indistinguishable from the erythrocyte high-activity form, whereas the other, also possessing high activity, was found only in the endometrial or uterine tissue. Present evidence suggests that the former isoenzyme originated from residual blood contaminating the tissue homogenates, and that a marked rise in the content of the latter isoenzyme accounts for the increase in rabbit endometrial carbonic anhydrase activity that previously has been observed in early pregnancy. 3. Minor forms of the erythrocyte isoenzymes, having a characteristic quantitative and electrophoretic relationship to one another, were occasionally produced during purification. 4. The actions were investigated of the inhibitors acetazolamide (5-acetamido-3,4-diazole-1-thia-2-sulphonamide), 1,1-dimethylaminonaphthalene-5-sulphonamide and ethoxyzolamide (6-ethoxybenzothiazole-2-sulphonamide) on the hydration of carbon dioxide and the hydrolysis of p-nitrophenyl acetate catalysed by the isoenzymes. 5. The low-activity erythrocyte isoenzyme was superior to the high-activity form as a catalyst of beta-naphthyl acetate hydrolysis.
D000818:Animals; D002245:Carbon Dioxide / Q000378:metabolism; D002257:Carbonic Anhydrase Inhibitors / Q000494:pharmacology; D002256:Carbonic Anhydrases / Q000032:analysis*; D002852:Chromatography, Ion Exchange; D004586:Electrophoresis; D004717:Endometrium / Q000201:enzymology; D004912:Erythrocytes / Q000201:enzymology*; D005260:Female; D007525:Isoelectric Focusing; D007527:Isoenzymes / Q000032:analysis*; D007700:Kinetics; D008970:Molecular Weight; D009596:Nitrophenols / Q000378:metabolism; D011247:Pregnancy; D011270:Pregnancy, Animal; D011817:Rabbits; D014599:Uterus / Q000201:enzymology*; D015032:Zinc / Q000032:analysis
D002257:Carbonic Anhydrase Inhibitors; D007527:Isoenzymes; D009596:Nitrophenols; D002245:Carbon Dioxide; D002256:Carbonic Anhydrases; D015032:Zinc