Cloning and expression of the bovine 11β-hydroxysteroid dehydrogenase type-2

The bovine 11 beta-hydroxysteroid dehydrogenase type 2 enzyme (11 beta-HSD-2) cDNA was cloned from three overlapping PCR fragments using primers based on the human and ovine 11 beta-HSD-2 cDNA sequences. Both cDNA ends were obtained by a modified RACE (Rapid Amplification of cDNA Ends) method. The bovine 11 beta-HSD-2 cDNA is 1878 bp long, excluding the poly(A) tail. It consists of a 5'-untranslated region of 133 bp, an open reading frame of 1215 bp and a 3'-untranslated region of 530 bp. Bovine 11 beta-HSD-2 cDNA is highly homologous to that of the sheep (92%) and less related to the human (67%), rabbit (65%), rat (52%) and mouse (45%) cDNA. The predicted bovine 11 beta-HSD-2 protein contains 404 amino acid residues with a calculated mol wt of 43,985. It is homologous to the sheep (98%) and human (88%) protein, and less related to that of the rabbit (76%), rat (80%) and mouse (77%). The cloned 11 beta-HSD-2 cDNA was transfected into CHOP cells and the enzymatic characteristics determined. The enzyme functions primarily as an oxidase, uses NAD(+) and is more active with corticosterone as a substrate than with cortisol or dexamethasone. It is expressed in high concentrations in kidney, adrenal and colon, and in small concentrations in liver, heart and lung. In conclusion, the 11 beta-HSD-2 enzyme of cattle is very similar to that of other species in its structure and enzymatic characteristics. (C) 2000 Elsevier Science Ltd. All rights reserved.