Biomedical subjects

Udo Oppermann

Publications and source records attributed to Udo Oppermann.

25 records · Page 2Linked to original sources

PubMed2003

Short-chain dehydrogenases/reductases (SDR): the 2002 update.

Short-chain dehydrogenases/reductases (SDR) form a large, functionally heterogeneous protein family presently with about 3000 primary and about 30 3D structures deposited in databases. Despite low sequence identities between different forms (about 15-30%), the 3D structures display highly similar alpha/beta folding patterns with a central beta-sheet, typical of the Rossmann-fold. Based on distinct sequence motifs functional assignments and classifications are possible, making it possible to build a general nomenclature system. Recent mutagenetic and structural studies considerably extend the knowledge on the general reaction mechanism, thereby establishing a catalytic tetrad of Asn-Ser-Tyr-Lys residues, which presumably form the framework for a proton relay system including the 2'-OH of the nicotinamide ribose, similar to the mechanism found in horse liver ADH. Based on their cellular functions, several SDR enzymes appear as possible and promising pharmacological targets with application areas spanning hormone-dependent cancer forms or metabolic diseases such as obesity and diabetes, and infectious diseases.

Crystallography, X-Ray↗

PubMed2003

Coenzyme-based functional assignments of short-chain dehydrogenases/reductases (SDRs).

Short-chain dehydrogenases/reductases (SDRs) are enzymes of great functional diversity. In spite of a residue identity of only 15-30%, the folds are conserved to a large extent, with specific sequence motifs detectable. We have developed an assignment scheme based on these motifs and detect five families. Only two of these were known before, called 'Classical' and 'Extended', but are now distinguished at a further level based on patterns of charged residues in the coenzyme-binding region, giving seven subfamilies of classical SDRs and three subfamilies of extended SDRs. Three further families are novel entities, denoted 'Intermediate', 'Divergent' and 'Complex', encompassing short-chain alcohol dehydrogenases, enoyl reductases and multifunctional enzymes, respectively. The assignment scheme was applied to the genomes of human, mouse, D. melanogaster, C. elegans, A. thaliana and S. cerevisiae. In the animal genomes, genes corresponding to the extended SDRs amount to around one quarter or less of the total number of SDR genes, while in those of A. thaliana and S. cerevisiae, the extended members constitute about 40% of the SDR forms. The NAD(H)-dependent SDRs are about equally many as the NADP(H)-dependent ones in human, mouse and plant, while the proportions of NAD(H)-dependent enzymes are much lower in fruit fly, worm and yeast. We also find that NADP(H) is the preferred coenzyme among most classical SDRs, while NAD(H) is that preferred among most extended SDRs.

Animals↗

PubMed2002

Comparative enzymology of 11 beta -hydroxysteroid dehydrogenase type 1 from glucocorticoid resistant (Guinea pig) versus sensitive (human) species.

Type 1 11 beta-hydroxysteroid dehydrogenase constitutes a prereceptor control mechanism through its ability to reduce dehydroglucocorticoids to the receptor ligands cortisol and corticosterone in vivo. We compared kinetic characteristics of the human and guinea pig 11 beta-hydroxysteroid dehydrogenase isozymes derived from species differing in glucocorticoid sensitivity. Both orthologs were successfully expressed as full-length enzymes in yeast and COS7 cells and as soluble transmembrane-deleted constructs in Escherichia coli. Both isozymes display Michaelis-Menten kinetics in intact cells and homogenates and show low apparent micromolar K(m) values in homogenates, which are lowered by approximately one order of magnitude in intact cells, allowing corticosteroid activation at physiological glucocorticoid levels. Recombinant soluble proteins were expressed and purified with high specific dehydrogenase and reductase activities, revealing several hundred-fold higher specificity constants than those reported earlier for the purified native enzyme. Importantly, these purified soluble enzymes also display a hyperbolic dependence of reaction velocity versus substrate concentration in 11-oxoreduction with K(m) values of 0.8 microm (human) and 0.6 microm (guinea pig), close to the values obtained from intact cells. Active site titration was carried out with the human enzyme using a novel inhibitor compound and reveals a fraction of 40-50% active sites/mol total enzyme. The kinetic data obtained argue against the involvement of 11 beta-hydroxysteroid dehydrogenase as a modulating factor for the glucocorticoid resistance observed in guinea pigs. Instead, the expression of 11 beta-hydroxysteroid dehydrogenase type 1 in the Zona glomerulosa of the guinea pig adrenal gland suggests a role of this enzyme in mineralocorticoid synthesis in this hypercortisolic species.

11-beta-Hydroxysteroid Dehydrogenase Type 2↗

PubMed2002

Critical residues for structure and catalysis in short-chain dehydrogenases/reductases.

Short-chain dehydrogenases/reductases form a large, evolutionarily old family of NAD(P)(H)-dependent enzymes with over 60 genes found in the human genome. Despite low levels of sequence identity (often 10-30%), the three-dimensional structures display a highly similar alpha/beta folding pattern. We have analyzed the role of several conserved residues regarding folding, stability, steady-state kinetics, and coenzyme binding using bacterial 3beta/17beta-hydroxysteroid dehydrogenase and selected mutants. Structure determination of the wild-type enzyme at 1.2-A resolution by x-ray crystallography and docking analysis was used to interpret the biochemical data. Enzyme kinetic data from mutagenetic replacements emphasize the critical role of residues Thr-12, Asp-60, Asn-86, Asn-87, and Ala-88 in coenzyme binding and catalysis. The data also demonstrate essential interactions of Asn-111 with active site residues. A general role of its side chain interactions for maintenance of the active site configuration to build up a proton relay system is proposed. This extends the previously recognized catalytic triad of Ser-Tyr-Lys residues to form a tetrad of Asn-Ser-Tyr-Lys in the majority of characterized short-chain dehydrogenases/reductase enzymes.

Amino Acid Sequence↗

PubMed2002

Short-chain dehydrogenases/reductases (SDRs).

Short-chain dehydrogenases/reductases (SDRs) are enzymes of great functional diversity. Even at sequence identities of typically only 15-30%, specific sequence motifs are detectable, reflecting common folding patterns. We have developed a functional assignment scheme based on these motifs and we find five families. Two of these families were known previously and are called 'classical' and 'extended' families, but they are now distinguished at a further level based on coenzyme specificities. This analysis gives seven subfamilies of classical SDRs and three subfamilies of extended SDRs. We find that NADP(H) is the preferred coenzyme among most classical SDRs, while NAD(H) is that preferred among most extended SDRs. Three families are novel entities, denoted 'intermediate', 'divergent' and 'complex', encompassing short-chain alcohol dehydrogenases, enoyl reductases and multifunctional enzymes, respectively. The assignment scheme was applied to the genomes of human, mouse, Drosophila melanogaster, Caenorhabditis elegans, Arabidopsis thaliana and Saccharomyces cerevisiae. In the animal genomes, the extended SDRs amount to around one quarter or less of the total number of SDRs, while in the A. thaliana and S. cerevisiae genomes, the extended members constitute about 40% of the SDR forms. The numbers of NAD(H)-dependent and NADP(H)-dependent SDRs are similar in human, mouse and plant, while the proportions of NAD(H)-dependent enzymes are much lower in fruit fly, worm and yeast. We show that, in spite of the great diversity of the SDR superfamily, the primary structure alone can be used for functional assignments and for predictions of coenzyme preference.

Amino Acid Motifs↗

PubMed2002

Short-chain dehydrogenase/reductase (SDR) relationships: a large family with eight clusters common to human, animal, and plant genomes.

The progress in genome characterizations has opened new routes for studying enzyme families. The availability of the human genome enabled us to delineate the large family of short-chain dehydrogenase/reductase (SDR) members. Although the human genome releases are not yet final, we have already found 63 members. We have also compared these SDR forms with those of three model organisms: Caenorhabditis elegans, Drosophila melanogaster, and Arabidopsis thaliana. We detect eight SDR ortholog clusters in a cross-genome comparison. Four of these clusters represent extended SDR forms, a subgroup found in all life forms. The other four are classical SDRs with activities involved in cellular differentiation and signalling. We also find 18 SDR genes that are present only in the human genome of the four genomes studied, reflecting enzyme forms specific to mammals. Close to half of these gene products represent steroid dehydrogenases, emphasizing the regulatory importance of these enzymes.

Animals↗

PubMed2002

Liver X receptors downregulate 11beta-hydroxysteroid dehydrogenase type 1 expression and activity.

11Beta-hydroxysteroid dehydrogenase type 1 (11beta-HSD-1) converts inactive corticosteroids into biologically active corticosteroids, thereby regulating the local concentration of active glucocorticoids, such as cortisol. 11beta-HSD-1 is particularly expressed in adipocytes and liver and appears to be causally linked to the development of type 2 diabetes and the metabolic syndrome. Liver X receptor (LXR)-alpha and -beta are nuclear oxysterol receptors whose key role in lipid metabolic regulation has recently been established. In this study, we show that treatment of adipocytes derived from 3T3-L1 cells and mouse embryonic fibroblasts in vitro with synthetic or natural LXR agonists decreases mRNA expression of 11beta-HSD-1 by approximately 50%, paralleled by a significant decline in 11beta-HSD-1 enzyme activity. Downregulation of 11beta-HSD-1 mRNA by LXRs started after a lag period of 8 h and required ongoing protein synthesis. Moreover, long-term per os treatment with a synthetic LXR agonist downregulated 11beta-HSD-1 mRNA levels by approximately 50% in brown adipose tissue and liver of wild-type but not of LXRalpha(-/-)beta(-/-) mice and was paralleled by downregulation of hepatic PEPCK expression. In conclusion, LXR ligands could mediate beneficial metabolic effects in insulin resistance syndromes including type 2 diabetes by interfering with peripheral glucocorticoid activation.

11-beta-Hydroxysteroid Dehydrogenase Type 1↗