THE HUMAN 11-BETA-HYDROXYSTEROID DEHYDROGENASE TYPE-II ENZYME - COMPARISONS WITH OTHER SPECIES AND LOCALIZATION TO THE DISTAL NEPHRON

Effective glucocorticoid inactivation is currently thought to be an in dispensable feature of mineralocorticoid target cells. The enzyme 11 b eta-hydroxysteroid dehydrogenase (11 beta-HSD) inactivates glucocortic oids and prevents them from binding to the non-selective mineralocorti coid receptor. In the kidney it is the NAD dependent high affinity iso form (11 beta-HSD2) which is thought to endow specificity on the recep tor. The recent cloning of the human, sheep and rabbit 11 beta-HSD2 en zymes permits a comparison of the enzyme from the three species. Human and rabbit enzymes are 87% identical and of similar length, while the human and sheep enzymes have only 75% identity. The last 12 residues in all three species were found to be highly divergent, but most of th e ovine dishomology can be accounted for by the deletion of a single n ucleotide toward the C-terminus of the protein resulting in a shift in reading frame generating a protein 27 residues longer than the human isoform. Numerous other deletions were also observed in this region of the sheep cDNA sequence. Furthermore, the rabbit cDNA also displayed a large degree of dishomology with the human sequence a short distance downstream from the termination codon. Conserved overlapping cytoplas mic translocation signals were observed in all three species, suggesti ng a topology whereby the enzyme is anchored into the endoplasmic reti culum by multiple hydrophobic regions in the N-terminus and the bulk o f the 11 beta-HSD2 peptide is sited in the cytoplasm. A polyclonal ant ibody generated against the C-terminus of human 11 beta-HSD2 was used to localize the enzyme within the kidney. A high level of immunoreacti vity was observed in distal tubules and collecting ducts, localizing t he enzyme to the same part of the nephron as the mineralocorticoid rec eptor. Moderate levels of staining were also seen in vascular smooth m uscle cells. These results support the notion that 11 beta-HSD2 is an autocrine protector of the mineralocorticoid receptor and that it play s an important role in cardiovascular homeostatic mechanisms.