Structure-function relationships of 3β-hydroxysteroid dehydrogenase: Contribution made by the molecular genetics of 3β-hydroxysteroid dehydrogenase deficiency

Morel, Yves; Mébarki, Farida; Rhéaume, Éric; Sánchez, Rocío; Forest, Maguelone G.; Simard, Jacques

doi:10.1016/s0039-128x(96)00178-x

Cited by 65 publications

(57 citation statements)

References 44 publications

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“…Many rare mutations in the HSD3B2 gene have been reported previously, mainly because they were observed in DNA obtained from patients suffering from classical HSD3B2 deficiency [7,33]. However, none of those mutations was observed during our resequencing studies.…”

Section: Human Hsd3b1 and Hsd3b2 Resequencingmentioning

confidence: 65%

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Human 3β-hydroxysteroid dehydrogenase types 1 and 2: Gene sequence variation and functional genomics

Wang

Salavaggione

Pelleymounter

et al. 2007

The Journal of Steroid Biochemistry and Molecular Biology

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The 3β-hydroxysteroid dehydrogenase/Δ 5 -Δ 4 isomerase isoenzymes 1 and 2 (HSD3B1 and HSD3B2) are membrane-bound enzymes that play essential roles in the biosynthesis of steroid hormones. Therefore, variation in the HSD3B1 and HSD3B2 genes might play a role in the pathophysiology of steroid hormone-related disease. We set out to systematically identify common polymorphisms and haplotypes in human HSD3B1 and HSD3B2. We identified 17 single nucleotide polymorphisms (SNPs) in HSD3B1 and 9 in HSD3B2 -the majority of which were not present in public databases -by resequencing human HSD3B1 and HSD3B2 using 240 DNA samples from four different ethnic groups (60 samples per group). Functional genomic studies of the 5 nonsynonymous cSNPs in HSD3B1 and the one observed in HSD3B2 showed that two of these polymorphisms resulted in significant decreases in the quantity of enzyme protein expressed. However, none of the 3 nonsynonymous SNPs located in areas encoding putative membrane-binding domains altered subcellular localization of the enzyme as determined by immunofluorescence microscopy. Finally, common variant haplotypes in the 5′-flanking regions of these genes showed significant cell linedependent variation in their ability to drive transcription. In aggregate, these results provide a basis for study of the possible role in human disease of common genetic variation in HSD3B1 and HSD3B2.

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Section: Human Hsd3b1 and Hsd3b2 Resequencingmentioning

confidence: 65%

“…Structural analysis and mutagenesis studies have indicated that HSD3B1 and HSD3B2 contain two characteristic Y-X-X-X-K sequence motifs that are located between Tyr154 and Lys158 and between Tyr 269 and Lys273 [7,33,35]. These motifs are thought to be associated with the catalytic sites of the enzymes.…”

Section: Human Hsd3b1 and Hsd3b2 Resequencingmentioning

confidence: 99%

Human 3β-hydroxysteroid dehydrogenase types 1 and 2: Gene sequence variation and functional genomics

Wang

Salavaggione

Pelleymounter

et al. 2007

The Journal of Steroid Biochemistry and Molecular Biology

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“…Despite the widespread distribution of these proteins and their large number, in only a few cases has an essential function been assigned to a single 3β-HSD enzyme. These exceptions include the human C 19/21 3β-HSD type II enzyme, which is required for the synthesis of all steroid hormone classes (25); the product of the Nsdh1 gene in the mouse, which catalyzes reactions in the terminal steps of cholesterol biosynthesis (26); the 3β-HSD enzyme encoded by the Sax1 gene of the plant Arabidopsis, which is required for brassinosteroid biosynthesis (27); and the presently described C 27 3β-HSD enzyme of bile acid biosynthesis. It is not yet clear why there are so many 3β-HSD enzymes and whether, as is the case with the four enzymes noted above, each member plays a unique metabolic role.…”

Section: Figurementioning

confidence: 99%

The bile acid synthetic gene 3β-hydroxy-Δ5-C27-steroid oxidoreductase is mutated in progressive intrahepatic cholestasis

Schwarz¹,

Wright²,

Davis³

et al. 2000

J. Clin. Invest.

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We used expression cloning to isolate cDNAs encoding a microsomal 3β-hydroxy-∆ 5 -C 27 -steroid oxidoreductase (C 27 3β-HSD) that is expressed predominantly in the liver. The predicted product shares 34% sequence identity with the C 19 and C 21 3β-HSD enzymes, which participate in steroid hormone metabolism. When transfected into cultured cells, the cloned C 27 3β-HSD cDNA encodes an enzyme that is active against four 7α-hydroxylated sterols, indicating that a single C 27 3β-HSD enzyme can participate in all known pathways of bile acid synthesis. The expressed enzyme did not metabolize several different C 19/21 steroids as substrates. The levels of hepatic C 27 3β-HSD mRNA in the mouse are not sexually dimorphic and do not change in response to dietary cholesterol or to changes in bile acid pool size. The corresponding human gene on chromosome 16p11.2-12 contains six exons and spans 3 kb of DNA, and we identified a 2-bp deletion in the C27 3β-HSD gene of a patient with neonatal progressive intrahepatic cholestasis. This mutation eliminates the activity of the enzyme in transfected cells. These findings establish the central role of C 27 3β-HSD in the biosynthesis of bile acids and provide molecular tools for the diagnosis of a third type of neonatal progressive intrahepatic cholestasis associated with impaired bile acid synthesis.

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“…A Tyr residue acts as the catalytic base, the hydroxyl group of Ser binds to the targeted oxo group to stabilize the substrate, and a Lys residue binds to the nicotinamide ribose and lowers the pKa of the phenolic hydroxyl group of the key Tyr to promote the proton transfer (Filling et al 2002). In a few species of 3 -HSD (macque, bovine, rat I, II and IV, guinea pig and chicken), Thr124 is positioned to perform the function of Ser124 (Simard et al 1996, Morel et al 1997). Our homology model of the three-dimensional structure of human 3 -HSD1 has been used to predict key amino acid residues involved in catalysis and coenzyme specificity (Thomas et al 2002(Thomas et al , 2003.…”

Section: Introductionmentioning

confidence: 99%

Serine 124 completes the Tyr, Lys and Ser triad responsible for the catalysis of human type 1 3beta-hydroxysteroid dehydrogenase

Thomas

Duax²,

Addlagatta³

et al. 2004

Journal of Molecular Endocrinology

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Human 3 -hydroxysteroid dehydrogenase/isomerase (3 -HSD) is a key steroidogenic enzyme that catalyzes the first step in the conversion of circulating dehydroepiandrosterone (DHEA), pregnenolone or 17 -hydroxypregenolone to produce the appropriate, active steroid hormone(s): estradiol, testosterone, progesterone, aldosterone or cortisol respectively. Our mutagenesis studies have identified Tyr154 and Lys158 as catalytic residues for the 3 -HSD reaction. Our three-dimensional homology model of 3 -HSD shows that Tyr154 and Lys158 are oriented near the 3 -hydroxyl group of the bound substrate steroid, and predicts that Ser123 or Ser124 completes a Tyr-Lys-Ser catalytic triad that operates in many other dehydrogenases. The S123A and S124A mutants of human type 1 3 -hydroxysteroid dehydrogenase/ isomerase (3 -HSD1) were created by PCR-based mutagenesis, expressed in insect cells using baculovirus and purified to homogeneity. The S124A mutant exhibits no 3 -HSD activity and has a K m value (83·6 µM) for the isomerase substrate that is threefold greater than that of wild-type 1 isomerase. In contrast, S123A has substantial 3 -HSD activity (DHEA K m =11·2 µM; k cat =0·8 min -1 ) and utilizes isomerase substrate, 5-androstene-3,17-dione, with a K m value (27·6 µM) that is almost identical to wild-type. The K m value (4·3 µM) of S124A for NADH as an allosteric activator of isomerase is similar to that of the wild-type 1 enzyme, indicating that Ser124 is not involved in cofactor binding. S123A utilizes NAD as a cofactor for 3 -HSD and NADH as the activator for isomerase with K m values that are similar to wild-type. The 3 -HSD activities of S123A and wild-type 3 -HSD increase by 2·7-fold when the pH is raised from 7·4 to the optimal pH 9·7, but S124A exhibits very low residual 3 -HSD activity that is pH-independent.These kinetic analyses strongly suggest that the Ser124 residue completes the catalytic triad for the 3 -HSD activity. Since there are 29 Ser residues in the primary structure of human 3 -HSD1, our homology model of the catalytic domain has been validated by this accurate prediction. A role for Ser124 in the binding of the isomerase substrate, which is the 3 -HSD product-steroid of the bifunctional enzyme protein, is also suggested. These observations further characterize the structure/function relationships of human 3 -HSD and bring us closer to the goal of selectively inhibiting the type 1 enzyme in placenta to control the timing of labor or in hormone-sensitive breast tumors to slow their growth.

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Structure-function relationships of 3β-hydroxysteroid dehydrogenase: Contribution made by the molecular genetics of 3β-hydroxysteroid dehydrogenase deficiency

Cited by 65 publications

References 44 publications

Human 3β-hydroxysteroid dehydrogenase types 1 and 2: Gene sequence variation and functional genomics

Human 3β-hydroxysteroid dehydrogenase types 1 and 2: Gene sequence variation and functional genomics

The bile acid synthetic gene 3β-hydroxy-Δ5-C27-steroid oxidoreductase is mutated in progressive intrahepatic cholestasis

Serine 124 completes the Tyr, Lys and Ser triad responsible for the catalysis of human type 1 3beta-hydroxysteroid dehydrogenase

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