Cloning and expression of human liver dehydroepiandrosterone sulphotransferase

Comer, K A; Falany, Josie L.; Falany, Charles N.

doi:10.1042/bj2890233

Cited by 118 publications

(73 citation statements)

References 23 publications

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“…Protein Purification-Human SULT DNA was codon optimized for E. coli (Mr. Gene, Regensburg, Germany) and inserted into a triple-tag pGEX-6P expression vector with an N-terminal His/GST/maltose-binding protein tag (13,14). The plasmid was transfected into E. coli (BL-21(DE3)), expressed, and purified as described previously (15).…”

Section: Methodsmentioning

confidence: 99%

High Accuracy in Silico Sulfotransferase Models

Cook

Wang

Falany

et al. 2013

Journal of Biological Chemistry

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Background: Human sulfotransferases (SULTs) regulate the bioactivities of hundreds of compounds in vivo. Results:The in silico models of SULTs developed here proved remarkably accurate in identifying SULT substrates in a 1,455-compound library containing all FDA-approved drugs. Conclusion: Highly accurate in silico SULT models are now available. Significance: The elision of mechanism and modeling can produce remarkably accurate in silico tools for the study of biology.

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Section: Methodsmentioning

confidence: 99%

High Accuracy in Silico Sulfotransferase Models

Cook

Wang

Falany

et al. 2013

Journal of Biological Chemistry

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“…A single isoform of DHEA-ST from human liver and adrenal tissues was confirmed by the expression and purification of the enzyme from these organs (5,6), molecular cloning studies (7) and the comparison study of the physical, kinetic, and immunological properties of liver and adrenal forms of the enzyme (8). Steroid sulfonation has been recognized as an important means for maintaining steroid hormone levels in their metabolism.…”

mentioning

confidence: 95%

Identifying Androsterone (ADT) as a Cognate Substrate for Human Dehydroepiandrosterone Sulfotransferase (DHEA-ST) Important for Steroid Homeostasis

Chang

Shi

Rehse

et al. 2004

Journal of Biological Chemistry

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In steroid biosynthesis, human dehydroepiandrosterone sulfotransferase (DHEA-ST) in the adrenals has been reported to catalyze the transfer of the sulfonate group from 3-phosphoadenosine-5-phosphosulfate to dehydroepiandrosterone (DHEA). DHEA and its sulfate play roles as steroid precursors; however, the role of the enzyme in the catabolism of androgens is poorly understood. Androsterone sulfate is clinically recognized as one of the major androgen metabolites found in urine. Here it is demonstrated that this enzyme recognizes androsterone (ADT) as a cognate substrate with similar kinetics but a 2-fold specificity and stronger substrate inhibition than DHEA. The structure of human DHEA-ST in complex with ADT has been solved at 2.7 Å resolution, confirming ADT recognition. Structural analysis has revealed the binding mode of ADT differs from that of DHEA, despite the similarity of the overall structure between the ADT and the DHEA binary complexes. Our results identify that this human enzyme is an ADT sulfotransferase as well as a DHEA sulfotransferase, implying an important role in steroid homeostasis for the adrenals and liver.Sulfonation is catalyzed by a family of sulfotransferases that conjugate a sulfonate group (SO 3 ) from 3Ј-phosphoadenosine-5Ј-phosphosulfate (PAPS) 1 to a hydroxyl group of the recipient molecule. With desulfation by sulfatases, sulfonation has been considered as one of the major enzymatic reactions in the metabolism not only of endogenous compounds and xenobiotics, but also of steroid hormones. In most cases, the transfer of the charged sulfonate moiety to an acceptor steroid decreases the biological activity of the steroid. Indeed, steroid sulfates resulting from this reaction are not capable of binding to or activating steroid receptors. In addition, the sulfonation reaction increases water solubility of steroids and thereby enhances their excretion into the urine and/or bile (1, 2).Human dehydroepiandrosterone sulfotransferase (DHEA-ST; SULT2A1; EC 2.8.2.2) was identified mainly from human liver and adrenals, using Northern blot analysis (3) and RT-PCR analysis (4). A single isoform of DHEA-ST from human liver and adrenal tissues was confirmed by the expression and purification of the enzyme from these organs (5, 6), molecular cloning studies (7) and the comparison study of the physical, kinetic, and immunological properties of liver and adrenal forms of the enzyme (8). Steroid sulfonation has been recognized as an important means for maintaining steroid hormone levels in their metabolism. In humans, dehydroepiandrosterone sulfate (DHEAS) is the most prodigious steroid precursor and one of the major secretory products of both adult and fetal adrenals. In the fetoplacental-maternal unit (the unique interdependence of fetus, placenta, and mother) shown in Scheme 1, DHEAS plays an important role as the major precursor for placental estrogen biosynthesis, thus maintaining pregnancy. A considerable amount of DHEAS is mainly produced from the fetal zone in the adrenal gland (9). Then DHEAS...

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“…The phenol SULTs, P-PST-1 (ST1A1) and M-PST (ST1A3), which have been localized in brain [4,6], are also expressed in the liver and\or intestine at high levels. Hydroxysteroid SULTs identified in rat brain [19,20] are also expressed in liver [21,22]. The functions of the SULTs in brain are poorly understood, although sulphation and inactivation of monoamine neurotransmitters and enkephalins have been proposed as potential roles for the phenol SULTs.…”

Section: Discussionmentioning

confidence: 99%

“…His-tagged hBR-STL protein was purified by Ni# + -affinity chromatography and electroelution for use as an antigen, and as a standard in the immunoblotting procedures. Initial attempts to raise polyclonal antibodies in rabbits using procedures that were effective for other cytosolic SULTs [11,[15][16][17] were unsuccessful. Also attempts to raise rabbit antibodies to hBR-STL by commercial laboratories were unsuccessful.…”

Section: Immunoblot Analysis Of Rbr-stl In Rat Brainmentioning

confidence: 99%

Molecular cloning and expression of novel sulphotransferase-like cDNAs from human and rat brain

Falany

Xie

Wang

et al. 2000

Biochemical Journal

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The sulphotransferase (SULT) gene family is involved with the conjugation of many small drugs, xenobiotics and endogenous compounds. In this report, we describe the cloning and expression of novel cDNAs from human and rat brain, which are structurally related to the SULTs. These cDNAs have been termed ‘brain sulphotransferase-like’ (BR-STL), because of their similarity to the SULTs and their selective expression in brain tissue. The proteins encoded by the human and rat BR-STL cDNAs (hBR-STL-1 and rBR-STL cDNA respectively), denoted as hBR-STL and rBR-STL, are 98% identical and 99% similar in sequence. The hBR-STL-1 cDNA contains an 852-nt open reading frame encoding a 284-amino-acid protein with a calculated molecular mass of 33083 Da. Northern-blot analyses of RNA isolated from eight human tissues indicate that the hBR-STL message is selectively expressed in brain. Characterization of different brain regions showed that message levels were highest in cortical brain regions. rBR-STL message levels were relatively low in brains of 1-day-old male and female rats, but increased to adult levels in RNA from 7-day-old rats, and remained at that level in adult animals. The hBR-STL and rBR-STL cDNAs were expressed in both Escherichia coli and Sf9 insect cells in the presence or absence of an N-terminal histidine-affinity tag (His-tag). Polyclonal antibodies were raised in chickens against purified His-tagged hBR-STL, and were used to detect the presence of rBR-STL in adult male and female rat brain cytosol. The high degree of sequence conservation, and the selective localization of the BR-STL message in brain, suggest an important function in the central nervous system.

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Cloning and expression of human liver dehydroepiandrosterone sulphotransferase

Cited by 118 publications

References 23 publications

High Accuracy in Silico Sulfotransferase Models

High Accuracy in Silico Sulfotransferase Models

Identifying Androsterone (ADT) as a Cognate Substrate for Human Dehydroepiandrosterone Sulfotransferase (DHEA-ST) Important for Steroid Homeostasis

Molecular cloning and expression of novel sulphotransferase-like cDNAs from human and rat brain

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