CLONING AND PRIMARY STRUCTURE OF MURINE BETA-HYDROXYSTEROID DEHYDROGENASE MICROSOMAL CARBONYL REDUCTASE

Screening of a mouse liver lambda gt 11 cDNA library with a rat liver 11 beta-hydroxysteroid dehydrogenase cDNA (11 beta-HSDr1A) and subsequ ent screening with an isolated mouse probe, resulted in the isolation and structure determination of a mouse cDNA encoding an amino acid seq uence which is very similar to human and rat 11 beta-hydroxysteroid de hydrogenases (78% and 86% similar, respectively), and also to other kn own vertebrate 11 beta-hydroxysteroid dehydrogenase structures. Open-r eading-frame analysis and the deduced amino acid sequence predict a pr otein with a molecular mass of 32.3 kDa which belongs to the superfami ly of the short-chain dehydrogenase proteins. The amino acid sequence contains two potential glycosylation sites. These data are in agreemen t with information on the glycoprotein character of the native enzyme. This kind of post-translational modification seems to be a determinin g factor concerning the equilibrium of the catalyzed 11 beta-dehydroge nation/11-oxo reduction step [Obeid, J., Curnow, K. M., Aisenberg, J. and White, P. C. (1993) Mol. Endocrinol. 7, 154-160; Agarwal, A. K., T usie-Luna, M. T., Monder, C. and White, P. C. (1990) Mol. Endocrinol. 4, 1827-1832]. After in vitro transcription/translation of the mouse c DNA, immunoprecipitation with anti-(microsomal carbonyl reductase) ser um and N-terminal sequence analysis of the purified protein confirms t he identity of microsomal 11 beta-hydroxysteroid dehydrogenase with th e previously described, microsomal-bound xenobiotic carbonyl reductase [Maser, E. and Bannenberg, G. (1994) Biochem. Pharmacol. 47, 1805-181 2], and points to an involvement of the 11 beta-HSD1A isoform in the r eductive phase-I metabolism of xenobiotic compounds, besides its endoc rinological functions. The alignment and comparison to other hydroxyst eroid dehydrogenase forms of the same protein superfamily allows the i dentification of important residues in the 11 beta-HSD primary structu re.