Isolation and sequence determination of a cDNA clone for rat peroxisomal urate oxidase: liver-specific expression in the rat.

摘要

Urate oxidase (UOxase; urate:oxygen oxidoreductase, EC 1.7.3.3), which catalyzes the oxidation of uric acid to allantoin, is present in most mammals but is absent in humans and certain primates. A cDNA clone for UOxase containing an insert of 1.3 kilobases (kb) was isolated from a lambda gt11 cDNA library prepared from rat liver mRNA. This recombinant clone with a 1283-nucleotide insert has sequence for 97% of the coding region together with 401 nucleotides of the 3'-untranslated region of the mRNA. The identity of UOxase cDNA clone was verified by analyzing the fusion protein, immunocytochemical localization with epitope-selected antibody, and hybrid-select translation analysis and by comparing sequences of four CNBr-cleaved peptides of the protein. Blot analysis revealed that the probe hybridizes to a single 1.5-kb mRNA species in the rat liver and a transplantable hepatocellular carcinoma. No UOxase mRNA was detected in 11 nonhepatic tissues of rat, suggesting tissue specificity of expression of this UOxase gene. Blot analysis of RNA from livers of rats treated with a peroxisome proliferator showed 2- to 3-fold increase in UOxase mRNA content, whereas the fatty acyl-CoA oxidase mRNA increased over 30-fold. Southern blot analysis of restriction enzyme digests of rat DNA suggests that there is a single copy of UOxase gene. Analysis of human genomic DNA revealed restriction fragments that are homologous to rat UOxase cDNA, although no UOxase mRNA was detected in human liver.