Structural and Functional Insights into Aldosterone Biosynthesis

The steroid hormone aldosterone synthesized by cytochrome P450 11B2 (CYP11B2, aldosterone synthase) regulates blood pressure and mineral homeostasis. For aldosterone synthesis, CYP11B2 requires the small iron/sulfur cluster protein adrenodoxin (Adx) which provides electrons for the reaction. Overproduction of aldosterone leads to primary aldosteronism, the most common form of secondary hypertension with very limited treatment options. Herein a structure/function approach was used to examine the role of adrenodoxin as allosteric modulator of aldosterone biosynthesis and to define the structural basis for CYP11B2 interaction with adrenodoxin. Studies revealed that adrenodoxin binding on the CYP11B2 surface increases the affinity to its physiological substrate 11-deoxycorticosterone in the buried active site up to 5.5-fold. Kinetic studies showed an increase of CYP11B2 turnover and a decrease in Km values at higher adrenodoxin concentrations. When performing inhibition studies, higher concentrations of adrenodoxin enhance the inhibitory effect of the antihypertensive drug osilodrostat. To elucidate the structural basis for aldosterone biosynthesis, an adrenodoxin/CYP11B2 fusion protein was generated to examine the protein/protein interaction. An X-ray structure of this complex with the breast cancer drug fadrozole was obtained in high resolution and identified the adrenodoxin binding site on the proximal face of CYP11B2 and specific electrostatic interactions. Overall, the studies presented here improve our understanding of the aldosterone biosynthetic system and the impact of adrenodoxin on CYP11B2 interaction with substrates and drugs. This information can be exploited for an orthogonal approach to treat primary aldosteronism and associated cardiac disease.