Structural and biochemical insights into 7β-hydroxysteroid dehydrogenase stereoselectivity

Area of Science:

• Biochemistry
• Structural Biology
• Enzymology

Background:

• Hydroxysteroid dehydrogenases (HSDs) are enzymes crucial for steroid metabolism.
• Their high stereo- and regio-selectivity makes them attractive biocatalysts for steroid modifications.
• Understanding HSD structure-function relationships is key to harnessing their catalytic potential.

Purpose of the Study:

• To elucidate the structural basis of substrate specificity and stereoselectivity in 7β-hydroxysteroid dehydrogenase (7β-HSD).
• To compare the structure of 7β-HSD with its α-selective counterpart to understand differing stereochemical outcomes.
• To investigate the role of specific structural elements, such as active-site loops and C-terminal extensions, in enzyme function.

Main Methods:

• X-ray crystallography was used to determine the three-dimensional structure of 7β-hydroxysteroid dehydrogenase from Collinsella aerofaciens.
• Comparative structural analysis was performed with known structures of 7α-hydroxysteroid dehydrogenase.
• Bioinformatic tools were employed to analyze active-site architecture and substrate-binding interactions.

Main Results:

• The crystal structure revealed a gating loop controlling active-site accessibility and elucidated the basis for NADP(+) cofactor specificity.
• The study identified structural determinants responsible for the β-selectivity of the enzyme.
• A C-terminal extension was found to reshape the substrate-binding site, potentially inverting substrate orientation compared to α-selective enzymes.

Conclusions:

• The structure of 7β-HSD provides insights into the molecular mechanisms underlying its catalytic activity and substrate selectivity.
• Structural comparison with 7α-HSD explains the opposing stereoselectivity observed between these enzymes.
• The findings offer a foundation for the rational design of HSDs as biocatalysts for specific steroid transformations.