Rat liver 3α-hydroxysteroid dehydrogenase

Penning, T.M.; Smithgall, Thomas E.; Askonas, Leslie J.; Sharp, Robert

doi:10.1016/0039-128x(86)90094-2

Cited by 39 publications

(29 citation statements)

References 26 publications

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“…Ps3␣HSD uses both NAD(H) and NADP(H) as a coenzyme. Such a character is called "dual pyridine nucleotide specificity" (31)(32)(33)(34). On the other hand, the structures of Ct3␣HSD that are incapable of accepting NADP(H) as a coenzyme were determined in different crystals, with and without NAD, by Grimm et al (8).…”

Section: Discussionmentioning

confidence: 99%

“…The side chain of this Asp 32 was hydrogen-bonded to 2Ј-and 3Ј-hydroxyl groups of the adenosine ribose in the same manner in both proteins. In the case of NADP(H), the 2Ј-phosphate group of the adenosine moiety would be electrostatically repelled by Asp 32 . These interactions are considered to cause the NAD(H) preference of Ps3␣HSD and Ct3␣HSD.…”

Section: Discussionmentioning

confidence: 99%

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Apo- and Holo-structures of 3α-Hydroxysteroid Dehydrogenase fromPseudomonassp. B-0831

et al. 2006

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Bacterial 3␣-hydroxysteroid dehydrogenase, which belongs to a short-chain dehydrogenase/reductase family and forms a dimer composed of two 26-kDa subunits, catalyzes the oxidoreduction of hydroxysteroids in a coenzyme-dependent manner. This enzyme also catalyzes the oxidoreduction of nonsteroid compounds that play an important role in xenobiotic metabolism of bacteria. We performed an x-ray analysis on the crystal of Ps3␣HSD, the enzyme from Pseudomonas sp. B-0831 complexed with NADH. The resulting crystal structure at 1.8 Å resolution showed that Ps3␣HSD exists as a structural heterodimer composed of apo-and holo-subunits. A distinct structural difference between them was found in the 185-207-amino acid region, where the structure in the apo-subunit is disordered whereas that in the holo-subunit consists of two ␣-helices. This fact proved that the NADH binding allows the helical structures to form the substrate binding pocket even in the absence of the substrate, although the region corresponds to the so-called "substrate-binding loop." The induction of ␣-helices in solution by the coenzyme binding was also confirmed by the CD experiment. In addition, the CD experiment revealed that the helixinducing ability of NADH is stronger than that of NAD. We discuss the negative cooperativity for the coenzyme binding, which is caused by the effect of the structural change transferred between the subunits of the heterodimer.Hydroxysteroid dehydrogenases are classified into three superfamilies as follows: aldo-keto reductase, long/mediumchain dehydrogenase/reductase, and short-chain dehydrogenase/reductase (SDR).2 Bacterial 3␣-hydroxysteroid dehydrogenase from Pseudomonas sp. B-0831 (Ps3␣HSD, EC 1.1.1.50) belonging to the SDR family forms a dimer composed of two 26-kDa subunits and catalyzes the oxidoreduction of steroid compounds as the reversible inter-conversion between 3␣-hydroxysteroid and 3-ketosteroid. In addition, Ps3␣HSD can also catalyze the oxidoreduction of nonsteroid compounds, such as p-nitrobenzaldehyde, and therefore is also named as 3␣-hydroxysteroid dehydrogenase/carbonyl reductase (3␣HSD/CR) (1). Bacteria are supposed to use this 3␣HSD/CR activity for a xenobiotic metabolism that is the first step in the consecutive detoxification process in bacterial organisms (2, 3). The enzymes belonging to the SDR family form a common tertiary structure of a dinucleotide-binding motif, popularly known as the Rossmann fold (4). These structures have the same architecture as the Ser-Tyr-Lys triad (SYK triad) as an active site and the same coenzyme-binding motif GXXXGXG at the N-terminal region. Although they have quite similar coenzyme-binding sites, there is a unique specificity for NAD(H) and/or NADP(H). A number of studies have been performed to explain the coenzyme specificity in SDR and illuminated that the Asp residue at the coenzyme-binding site determines the preference of NAD(H). The switch of the coenzyme preference from NADP(H) to NAD(H) has been achieved by the mutation of this Asp residue (5). We have stud...

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Apo- and Holo-structures of 3α-Hydroxysteroid Dehydrogenase fromPseudomonassp. B-0831

et al. 2006

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“…1 To whom correspondence should be addressed (e-mail penning!pharm.med.upenn.edu). 2 ized 17β-oestradiol to oestrone, and 20α-hydroxyprogesterone to progesterone. Discrete tissue distribution of these AKR1C enzymes was observed using isoform-specific reverse transcriptase-PCR.…”

Section: Introductionmentioning

confidence: 99%

“…In the liver, 3α-HSDs work in concert with 3-oxo-5α-steroid-4-dehydrogenase (5α-reductase ; EC 1.3.99.5) and 5β-reductase to convert 5α\5β-dihydrosteroids into 5α\5β-tetrahydrosteroids [1,2]. In this manner they catalyse the second step Abbreviations used : AKR, aldo-keto reductase ; allopregnanolone, 3α-hydroxy-5α-pregnan-20-one ; 3α-androstanediol, 5α-androstane-3α,17β-diol ; androsterone, 3α-hydroxy-5α-androstan-17-one ; 5α-DHT, 5α-dihydrotestosterone (17β-hydroxy-5α-androstan-3-one) ; 5α-dihydroprogesterone, 5α-pregnane-3,20-dione ; GABA, γ-aminobutyric acid ; 3α-HSD, 3α-hydroxysteroid dehydrogenase ; 5α-reductase, 3-oxo-5α-steroid-4-dehydrogenase ; 20α-hydroxyprogesterone, 20α-hydroxy-pregn-4-ene-3,20-dione ; 3h-UTR, 3h-untranslated region ; RT, reverse transcriptase ; poly(A) + RNA, polyadenylated RNA.…”

Section: Introductionmentioning

confidence: 99%

Human 3α-hydroxysteroid dehydrogenase isoforms (AKR1C1–AKR1C4) of the aldo-keto reductase superfamily: functional plasticity and tissue distribution reveals roles in the inactivation and formation of male and female sex hormones

Penning

Burczynski

Jez

et al. 2000

Biochemical Journal

385

222

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The kinetic parameters, steroid substrate specificity and identities of reaction products were determined for four homogeneous recombinant human 3α-hydroxysteroid dehydrogenase (3α-HSD) isoforms of the aldo-keto reductase (AKR) superfamily. The enzymes correspond to type 1 3α-HSD (AKR1C4), type 2 3α(17β)-HSD (AKR1C3), type 3 3α-HSD (AKR1C2) and 20α(3α)-HSD (AKR1C1), and share at least 84 % amino acid sequence identity. All enzymes acted as NAD(P)(H)-dependent 3-, 17-and 20-ketosteroid reductases and as 3α-, 17β-and 20α-hydroxysteroid oxidases. The functional plasticity of these isoforms highlights their ability to modulate the levels of active androgens, oestrogens and progestins. Salient features were that AKR1C4 was the most catalytically efficient, with k cat \K m values for substrates that exceeded those obtained with other isoforms by 10-30-fold. In the reduction direction, all isoforms inactivated 5α-dihydrotestosterone (17β-hydroxy-5α-androstan-3-one ; 5α-DHT) to yield 5α-androstane-3α,17β-diol (3α-androstanediol). However, only AKR1C3 reduced ∆%-androstene-3,17-dione to produce significant amounts of testosterone. All isoforms reduced oestrone to 17β-oestradiol, and progesterone to 20α-hydroxypregn-4-ene-3,20-dione (20α-hydroxyprogesterone). In the oxidation direction, only AKR1C2 converted 3α-androstanediol to the active hormone 5α-DHT. AKR1C3 and AKR1C4 oxidized testosterone to ∆%-androstene-3,17-dione. All isoforms oxid-

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“…The results suggest that microsomal enzymes other than P450 enzymes were involved in ILCA formation. Epimerization of the hydroxyl group on steroid molecules is most often catalyzed by hepatic hydroxysteroid dehydrogenase and steroid oxidoreductase enzymes and is not a common P450-mediated reaction (Penning et al, 1986). A possible mechanism for the formation of ILCA is shown in Fig.…”

Section: Effect Of Sex and Treatment With P450 Inducers On Lithocholimentioning

confidence: 99%

Biotransformation of Lithocholic Acid by Rat Hepatic Microsomes: Metabolite Analysis by Liquid Chromatography/Mass Spectrometry

Deo

Bandiera²

2007

Drug Metab Dispos

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ABSTRACT:Lithocholic acid is a lipid-soluble hepatotoxic bile acid that accumulates in the liver during cholestasis. A potential detoxification pathway for lithocholic acid involves hydroxylation by hepatic cytochrome P450 (P450) enzymes. The purpose of the present study was to identify the hepatic microsomal metabolites of lithocholic acid by liquid chromatography/mass spectrometry and to determine the P450 enzymes involved. Incubation of lithocholic acid with rat hepatic microsomes and NADPH produced murideoxycholic acid (MDCA), isolithocholic acid (ILCA), and 3-keto-5␤-cholanic acid (3KCA) as major metabolites and 6-ketolithocholic acid and ursodeoxycholic acid as minor metabolites. Experiments with hepatic microsomes prepared from rats pretreated with P450 inducers and with inhibitory antibodies indicated that CYP2C and CYP3A enzymes contribute to microsomal MDCA formation. Results obtained with a panel of recombinant P450 enzymes and CYP2D6 antiserum showed that CYP2D1 can also catalyze MDCA formation. Similar experimental evidence revealed that formation of 3KCA was mediated primarily by CYP3A enzymes. ILCA formation appeared to be catalyzed by a distinct pathway mediated largely by microsomal non-P450 enzymes. Based on the results obtained using lithocholic acid and 3KCA as substrates, a mechanism for the formation of ILCA involving a geminal diol intermediate is outlined. In conclusion, lithocholic acid was extensively metabolized by multiple P450 enzymes with the predominant biotransformation pathway being hydroxylation at the 6␤-position. This study provides an insight into possible routes of detoxification of lithocholic acid.

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Rat liver 3α-hydroxysteroid dehydrogenase

Cited by 39 publications

References 26 publications

Apo- and Holo-structures of 3α-Hydroxysteroid Dehydrogenase fromPseudomonassp. B-0831

Apo- and Holo-structures of 3α-Hydroxysteroid Dehydrogenase fromPseudomonassp. B-0831

Human 3α-hydroxysteroid dehydrogenase isoforms (AKR1C1–AKR1C4) of the aldo-keto reductase superfamily: functional plasticity and tissue distribution reveals roles in the inactivation and formation of male and female sex hormones

Biotransformation of Lithocholic Acid by Rat Hepatic Microsomes: Metabolite Analysis by Liquid Chromatography/Mass Spectrometry

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