Satoko Fujiwara

Evolution of urate-degrading enzymes in animal peroxisomes.

The end product of purine metabolism varies from species to species. The degradation of purines to urate is common to all animal species, but the degradation of urate is much less complete in higher animals. The comparison of subcellular distribution, intraperoxisomal localization forms, molecular structures, and some other properties of urate-degrading enzymes (urate oxidase, allantoinase, and allantoicase) among animals is described. Liver urate oxidase (uricase) is located in the peroxisomes in all animals with urate oxidase. On the basis of the comparison of intraperoxisomal localization forms, mol wt, and solubility of liver urate oxidase among animals, it is suggested that amphibian urate oxidase is a transition form in the evolution of aquatic animals to land animals. Allantoinase and allantoicase are different proteins in fish liver, but the two enzymes form a complex in amphibian liver. The subcellular localization of allantoinase and allantoicase varies among fishes. Hepatic allantoinase is located both in the peroxisomes and in the cytosol in saltwater fishes, and only in the cytosol in freshwater fishes. Hepatic allantoicase is located on the outer surface of the, peroxisomal membrane in the mackerel group and in the peroxisomal matrix in the sardine group. Amphibian hepatic allantoinase-allantoicase complex is probably located in the mitochondria. On the basis of previous data, changes of allantoinase and allantoicase in molecular structure and intracellular localization during animal evolution may be as follows: Fish liver allantoinase is a single peptide with a mol wt of 54,000, and is located both in the peroxisomes and in the cytosol, or only in the cytosol. Fish liver allantoicase consists of two identical subunits with a mol wt of 48,000, and is located in the peroxisomal matrix or on the outer surface of the peroxisomal membrane. The evolution of fishes to amphibia resulted in the dissociation of allantoicase into subunits, and in the association of allantoinase with the subunit of allantoicase. This amphibian enzyme was lost by further evolution.

Comparison of intraperoxisomal localization form and properties of amphibian (Rana catesbeiana) uricase with those of other animal uricases

Liver uricase of bull frog (Rana catesbeiana) was present as the soluble form in the peroxisomal matrix and consisted of four identical subunits with a molecular weight of 30,000. These properties were identical with those of fish liver uricase but differed from mammalian liver uricase. Purified uricase from the frog liver was insoluble in hypertonic, hypotonic and detergent solutions at pH 6-9. This insolubility was the same as mammalian liver uricase but differed from fish liver uricase; fish uricase was soluble in these solutions. The frog liver uricase did not cross-react immunologically with both uricases of fish and mammalian liver. An immunological cross-reactivity of liver uricase was observed among amphibia.

Subcellular distribution of hepatic allantoinase varies among fishes

Abstract 1. 1. Intracellular and intraperoxisomal localization of hepatic allantoinase was examined by sucrose density gradient centrifugation with thirteen different fishes. 2. 2. On the basis of the localization of allantoinase, fishes were found to be classified into two groups. 3. 3. In group 1, allantoinase was located both in the peroxisomal soluble matrix and in the cytosol. In group 2, allantoinase was located only in the cytosol.

Insoluble uricase in liver peroxisomes of old world monkeys

1. Subcellular localization form and properties of liver uricase of Macaca fascicularis were examined. 2. Uricase was present as the insoluble form in the peroxisomal core. 3. Evidence was obtained to show that the peroxisomal core is uricase itself. 4. The number and mol. wts of the subunits of the enzyme were identical to those of rat liver uricase. 5. The same results were also obtained for liver uricase of Macaca fuscata.

Purification and characterization of peroxisomal apo and holo alanine:glyoxylate aminotransferase from bird liver

Alanine:glyoxylate aminotransferase has been reported to be present as the apo enzyme in the peroxisomes and as the holo enzyme in the mitochondria in chick (white leghorn) embryonic liver. However, surprisingly, birds were found to be classified into two groups on the basis of intraperoxisomal forms of liver alanine:glyoxylate aminotransferase. In the peroxisomes, the enzyme was present as the holo form in group 1 (pigeon, sparrow, Java sparrow, Australian budgerigar, canary, goose, and duck), and as the apo form in group 2 (white leghorn, bantam, pheasant, and Japanese mannikin). In the mitochondria, the enzyme was present as the holo form in both groups. The peroxisomal holo enzyme was purified from pigeon liver, and the peroxisomal apo enzyme from chicken (white leghorn) liver. The pigeon holo enzyme was composed of two identical subunits with a molecular weight of about 45,000, whereas the chicken apo enzyme was a single peptide with the same molecular weight as the subunit of the pigeon enzyme. The peroxisomal holo enzyme of pigeon liver was not immunologically cross-reactive with the peroxisomal apo enzyme of chicken liver, the mitochondrial holo enzymes from pigeon and chicken liver, and mammalian alanine:glyoxylate aminotransferases 1 and 2. The mitochondrial holo enzymes from both pigeon and chicken liver had molecular weights of about 200,000 with four identical subunits and were cross-reactive with mammalian alanine:glyoxylate aminotransferase 2 but not with mammalian alanine:glyoxylate aminotransferase 1.

Alanine:glyoxylate aminotransferase is present as the apoenzyme in the peroxisomes of chicken kidney

The subcellular distribution of alanine:glyoxylate aminotransferase in chicken kidney was examined by centrifugation in a sucrose density gradient. The enzyme was found to be present as the apoform in the peroxisomes and as the holoform in the mitochondria. Alanine:glyoxylate aminotransferase in different mammalian kidneys were all present as the holoenzyme in the mitochondrial and soluble fractions.

Is the guinea-pig (Cavia porcellus) a rodent ?

Abstract The Rodentia are traditionally divided into three extant suborders: the Sciuromorpha, the Myomorpha and the Hystricomorpha. The guinea-pig (Cavia porcellus) has been classified as a New World hystricomorph rodent. On the basis of the analysis of our previous studies on mammalian alanine:glyoxylate aminotransferase 1, we propose that the classification of the guinea-pig as a rodent is not correct.

Identity of alanine:glyoxylate aminotransferase with alanine:2-oxoglutarate aminotransferase in rat liver cytosol.

Rat liver soluble fraction contained 3 forms of alanine: glyoxylate aminotransferase. One with a pI of 5.2 and an Mr of approx. 110,000 was found to be identical with cytosolic alanine:2-oxoglutarate aminotransferase. The pI 6.0 enzyme with an Mr of approx. 220,000 was suggested to be from broken mitochondrial alanine:glyoxylate aminotransferase 2 and the pI 8.0 enzyme with an Mr of approx. 80,000 enzyme from broken peroxisomal and mitochondrial alanine:glyoxylate aminotransferase 1. These results suggest that the cytosolic alanine: glyoxylate aminotransferase activity is due to cytosolic alanine: 2-oxoglutarate aminotransferase.

Intraperoxisomal form of alanine:glyoxylate aminotransferase in the peroxisomes of bird kidney

Alanine:glyoxylate aminotransferase has been reported to be present as the apo form in the peroxisomes and as the holo form in the mitochondria in chicken kidney. In contrast, the enzyme was found to be present as the holo form both in the peroxisomes and in the mitochondria in pigeon kidney, suggesting that birds are classified into two groups on the basis of intraperoxisomal form of kidney alanine:glyoxylate aminotransferase. In the kidney, the pigeon peroxisomal holo enzyme did not cross-react immunologically with the chicken peroxisomal apo enzyme.

Enzymatic and immunological comparison of alanine: glyoxylate aminotransferases from different fish and mammalian livers

Alanine:glyoxylate aminotransferase was highly purified and characterized from mackerel liver. The purified enzyme had a mol. wt of approx. 200,000 with four identical subunits. It was specific for L-alanine and L-serine with glyoxylate and for L-serine with pyruvate as amino acceptor. The mackerel enzyme was similar to partially purified alanine:glyoxylate aminotransferases from other fish (sardine, gopher gray rock cod and yellow mackerel) liver with respect to mol. wts and substrate specificity. These fish enzymes were similar to mammalian liver alanine:glyoxylate aminotransferases 1 in substrate specificity and to mammalian alanine:glyoxylate aminotransferases 2 in mol. wts. An immunological cross-reactivity of hepatic alanine:glyoxylate aminotransferase was observed between mackerel and other fishes but not between mackerel and mammals.