Expression of human kidney 11β-hydroxysteroid dehydrogenase (11-HSD2) in bacteria

B. Scott Nunez, Tomoatsu Mune, Perrin C. White

Research output: Contribution to journalArticle

5 Scopus citations

Abstract

The kidney isozyme of 11β-hydroxysteroid dehydrogenase (11-HSD2) protects the mineralocorticoid receptor from spurious activation by glucocorticoids. To explore structure-function relationships, human 11-HSD2 cDNA was subcloned into the bacterial expression vector, pET25b. E. coli transformed with wild-type cDNA produced active enzyme that retained biochemical characteristics of the native protein. The addition of 6 histidine residues to the C-terminus of the wild-type enzyme (11-HSD2/His) increased activity 2-fold. Whereas wild-type activity was almost completely sedimented following 100,000 g centrifugation, 10-30% of total activity of 11-HSD2/His remained in the supernatant. The 11-HSD2 isozyme normally contains three N-terminal hydrophobic domains. Mutant 11-HSD2/His possessing a single hydrophobic domain retained partial activity, but elimination of all domains inactivated the enzyme. Thus, the N-terminal hydrophobic domains are essential for complete activity of 11-HSD2 but association with an intact cell membrane is not.

Original languageEnglish (US)
Pages (from-to)652-656
Number of pages5
JournalBiochemical and Biophysical Research Communications
Volume255
Issue number3
DOIs
StatePublished - Feb 24 1999

ASJC Scopus subject areas

  • Biophysics
  • Biochemistry
  • Molecular Biology
  • Cell Biology

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