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Short-chain dehydrogenases/reductases (SDR) constitute a large family of enzymes found in all forms of life. Despite a low level of sequence identity, the three-dimensional structures determined display a nearly superimposable alpha/beta folding pattern. We identified a conserved asparagine residue located within strand betaF and analyzed its role in the short-chain dehydrogenase/reductase architecture. Mutagenetic replacement of Asn179 by Ala in bacterial 3beta/17beta-hydroxysteroid dehydrogenase yields a folded, but enzymatically inactive enzyme, which is significantly more resistant to denaturation by guanidinium hydrochloride. Crystallographic analysis of the wild-type enzyme at 1.2-A resolution reveals a hydrogen bonding network, including a buried and well-ordered water molecule connecting strands betaE to betaF, a common feature found in 16 of 21 known three-dimensional structures of the family. Based on these results, we hypothesize that in mammalian 11beta-hydroxysteroid dehydrogenase the essential Asn-linked glycosylation site, which corresponds to the conserved segment, displays similar structural features and has a central role to maintain the SDR scaffold.

Original publication

DOI

10.1006/bbrc.2001.6032

Type

Journal article

Journal

Biochem Biophys Res Commun

Publication Date

07/12/2001

Volume

289

Pages

712 - 717

Keywords

11-beta-Hydroxysteroid Dehydrogenase Type 1, 17-Hydroxysteroid Dehydrogenases, Amino Acid Sequence, Asparagine, Circular Dichroism, Conserved Sequence, Glycosylation, Guanidine, Hydroxysteroid Dehydrogenases, Kinetics, Models, Molecular, Molecular Sequence Data, Mutagenesis, Site-Directed, Oxidoreductases, Protein Denaturation, Protein Folding, Sequence Homology, Amino Acid