11 beta-hydroxysteroid dehydrogenase-2 is a high affinity corticosterone-binding protein

Area of Science:

• Endocrinology
• Molecular Biology
• Renal Physiology

Background:

• Kidneys possess mineralocorticoid and glucocorticoid receptors.
• A third corticosteroid binding site, Type III, exhibits high affinity for corticosterone but not aldosterone or synthetic glucocorticoids.
• Similarities in steroid specificity suggested a link to 11 beta-hydroxysteroid dehydrogenase type 2 (11 beta-HSD2).

Purpose of the Study:

• To express recombinant rabbit 11 beta-HSD2 in mammalian cells.
• To determine if transfected cells acquire NAD-dependent 11 beta-HSD2 activity and high-affinity corticosterone binding sites.

Main Methods:

• Stable transfection of Chinese Hamster Ovary (CHO) cells with rabbit 11 beta-HSD2.
• Assay of NAD-dependent 11 beta-HSD2 activity.
• Measurement of corticosterone binding site affinity and specificity using radioligand competition assays.

Main Results:

• Transfected CHO cells exhibited high, NAD-dependent, unidirectional 11 beta-HSD2 activity.
• Acquisition of numerous corticosterone-specific binding sites (1.21 +/- 0.3 x 10^6) with a Kd of 25 +/- 8 nM.
• Binding specificity mirrored 11 beta-HSD2: corticosterone >> 11-hydroxyprogesterone > carbenoxolone > 11 dehydrocorticosterone > cortisol > progesterone ~ DOC >>> DEX > RU 28362 - aldosterone.
• No binding observed for glucocorticoid receptor (GR) or mineralocorticoid receptor (MR) agonists.

Conclusions:

• The Type III binding protein is identified as 11 beta-HSD2.
• 11 beta-HSD2 possesses high-affinity corticosterone binding sites.
• This enzyme plays a key role in regulating mineralocorticoid and glucocorticoid receptor activity in the kidney.