DHRS7 Dotaz Zobrazit nápovědu
The metabolism of steroids and retinoids has been studied in detail for a long time, as these compounds are involved in a broad spectrum of physiological processes. Many enzymes participating in the conversion of such compounds are members of the short-chain dehydrogenase/reductase (SDR) superfamily. Despite great effort, there still remain a number of poorly characterized SDR proteins. According to various bioinformatics predictions, many of these proteins may play a role in the metabolism of steroids and retinoids. Dehydrogenase/reductase (SDR family) member 7 (DHRS7) is one such protein. In a previous study, we determined DHRS7 to be an integral membrane protein of the endoplasmic reticulum facing the lumen which has shown at least in vitro NADPH-dependent reducing activity toward several eobiotics and xenobiotics bearing a carbonyl moiety. In the present paper pure DHRS7 was used for a more detailed study of both substrate screening and an analysis of kinetics parameters of the physiologically important substrates androstene-3,17-dione, cortisone and all-trans-retinal. Expression patterns of DHRS7 at the mRNA as well as protein level were determined in a panel of various human tissue samples, a procedure that has enabled the first estimation of the possible biological function of this enzyme. DHRS7 is expressed in tissues such as prostate, adrenal glands, liver or intestine, where its activity could be well exploited. Preliminary indications show that DHRS7 exhibits dual substrate specificity recognizing not only steroids but also retinoids as potential substrates and could be important in the metabolism of these signalling molecules.
- MeSH
- androstendion metabolismus MeSH
- cirkulární dichroismus MeSH
- fylogeneze MeSH
- kinetika MeSH
- kortison metabolismus MeSH
- lidé MeSH
- oxidoreduktasy chemie genetika metabolismus MeSH
- regulace genové exprese enzymů MeSH
- retinaldehyd metabolismus MeSH
- steroidy metabolismus MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Dehydrogenase/reductase SDR family member 7 (DHRS7, SDR34C1, retSDR4) is one of the many endoplasmic reticulum bound members of the SDR superfamily. Preliminary results indicate its potential significance in human metabolism. DHRS7 containing TEV-cleavable His10 and FLAG-tag expressed in the Sf9 cell line was solubilised, purified, and reconstituted into liposomes to enable the improved characterisation of this enzyme in the future. Igepal CA-630 was determined to be the best detergent for the solubilisation process. The solubilised DHRS7 was purified using affinity chromatography, and the purified enzyme was subjected to TEV cleavage of the affinity tags and then repurified using subtractive Ni-IMAC. The cleaved and uncleaved versions of DHRS7 were successfully reconstituted into liposomes. In addition, using tobacco specific carcinogen 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) as the substrate, the cleaved liposomal DHRS7 was found to be inactive, whereas the pure and uncleaved liposomal DHRS7 were confirmed as enzymes, which reduce carbonyl group of the substrates.
- MeSH
- buněčná membrána MeSH
- lidé MeSH
- membránové proteiny chemie genetika izolace a purifikace metabolismus MeSH
- oxidoreduktasy chemie genetika izolace a purifikace metabolismus MeSH
- rekombinantní proteiny chemie genetika izolace a purifikace metabolismus MeSH
- Sf9 buňky MeSH
- Spodoptera MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Human DHRS7 (SDR34C1) is one of insufficiently described enzymes of the short-chain dehydrogenase/reductase superfamily. The members of this superfamily often play an important pato/physiological role in the human body, participating in the metabolism of diverse substrates (e.g. retinoids, steroids, xenobiotics). A systematic approach to the identification of novel, physiological substrates of DHRS7 based on a combination of homology modeling, structure-based virtual screening and experimental evaluation has been used. Three novel substrates of DHRS7 (dihydrotestosterone, benzil and 4,4'-dimetylbenzil) have been described.
- Publikační typ
- abstrakt z konference MeSH
- Publikační typ
- abstrakt z konference MeSH
Dehydrogenase/reductase (SDR family) member 7 (DHRS7, retSDR4, SDR34C1) is a previously uncharacterized member of the short-chain dehydrogenase/reductase (SDR) superfamily. While human SDR members are known to play an important role in various (patho)biochemical pathways including intermediary metabolism and biotransformation of xenobiotics, only 20% of them are considered to be well characterized. Based on phylogenetic tree and SDR sequence clusters analysis DHRS7 is a close relative to well-known SDR member 11β-hydroxysteroid dehydrogenase 1 (11β-HSD1) that participates in metabolism of endogenous and xenobiotic substances with carbonyl group. The aim of present study is to determine the basic biochemical properties of DHRS7 and its possible involvement in metabolism of substrates with carbonyl group. For the first time the computational predictions of this membrane protein and membrane topology were experimentally confirmed. DHRS7 has been demonstrated to be an integral protein facing the lumen of the endoplasmic reticulum with lack of posttranscriptional glycosylation modification. Subsequently, NADP(H) cofactor preference and enzymatic reducing activity of DHRS7 was determined towards endogenous substrates with a steroid structure (cortisone, 4-androstene-3,17-dion) and also toward relevant exogenous substances bearing a carbonyl group harmful to human health (1,2-naphtoquinone, 9,10-phenantrenequinone). In addition to 11β-HSD1, DHRS7 is another enzyme from SDR superfamily that have been proved, at least in vitro, to contribute to the metabolism of xenobiotics with carbonyl group.
- MeSH
- 11-beta-hydroxysteroiddehydrogenasa typ 1 chemie metabolismus MeSH
- benzaldehydy metabolismus MeSH
- fluorescenční protilátková technika MeSH
- intracelulární membrány metabolismus MeSH
- izoenzymy chemie metabolismus MeSH
- jaterní mikrozomy enzymologie MeSH
- kinetika MeSH
- lidé MeSH
- molekulární sekvence - údaje MeSH
- NAD metabolismus MeSH
- NADP metabolismus MeSH
- nitrosaminy chemie metabolismus MeSH
- oxidoreduktasy chemie metabolismus MeSH
- sekvence aminokyselin MeSH
- Sf9 buňky MeSH
- spektrofotometrie MeSH
- substrátová specifita MeSH
- ultracentrifugace MeSH
- western blotting MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
