The enzyme 11β-hydroxysteroid dehydrogenase (11βHSD) is thought to protect the non-selective mineralocorticoid receptor from occupation by glucocorticoids, and to modulate access of glucocorticoids to glucocorticoid receptors resulting in protection of the fetus and gonads. A ubiquitous low affinity NADP+ dependent enzyme (11βHSD1) and a tissue specific, high affinity NAD+ dependent form (11βHSD2) of 11βHSD exist. We now report the isolation of a cDNA coding for human 11βHSD2. The new isoform is NAD+ dependent, exclusively dehydrogenase in directionality, inhibited by glycyrrhetinic acid and metabolizes the synthetic glucocorticoid dexamethasone; it displays K_m values for corticosterone and cortisol of 5.1 nM and 47 nM, respectively. Sequence alignment shows that 11βHSD2 shares 35% identity with 17βHSD2, but is only 14% identical with 11βHSD1. The 11βHSD2 gene is highly expressed in kidney, colon, pancreas and placenta and the message is also present in the ovary, prostate and testis. These data suggest that 11βHSD2 plays an important role in modulating mineralocorticoid and glucocorticoid receptor occupancy by glucocorticoids.