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>Isolation and Characterization of a Stereospecific 3β-Hydroxysteroid Sulfotransferase (Pregnenolone Sulfotransferase) cDNA
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Isolation and Characterization of a Stereospecific 3β-Hydroxysteroid Sulfotransferase (Pregnenolone Sulfotransferase) cDNA
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机译:Isolation and Characterization of a Stereospecific 3β-Hydroxysteroid Sulfotransferase (Pregnenolone Sulfotransferase) cDNA
ABSTRACTIn contrast to humans, who possess a hydroxysteroid sulfotransferase (HSST), namely, DHEA sulfotransferase (DHEA-ST), that displays broad substrate specificities, HSSTs of the guinea pig show a high substrate stereoselectivity, as shown by the recent cloning of a chiral-specific 3α-hydroxysteroid sulfotransferase. Herein, we report the cloning and expression of the substrate and chiral-specific pregnenolone sulfotransferase (PREG-ST). Transfection of the pCMV expression vector containing PREG-ST cDNA in transformed human embryonal kidney (293) cells showed that the expressed enzyme selectively catalyzes the 3β-hydroxysteroid substrate. It converts pregnenolone to pregnenolone sulfate most efficiently, whereas dehydroepiandrosterone and epiandrosterone were transformed at a much lower rate, and androsterone, a 3α-hydroxysteroid, was not significantly metabolized (30-fold lower). Thus, the enzyme was identified as pregnenolone sulfotransferase. DNA analysis predicts a protein of 287 amino acids with a calculated molecular mass of 34,199 daltons. Alignment of the amino acid sequence with other sulfotransferases indicated that guinea pig pregnenolone sulfotransferase shares 75 and 80 homology with human DHEA sulfotransferase and rat hydroxysteroid dehydrogenase, respectively. RNA blot analysis using guinea pig liver, intestine, adrenal, kidney, epididymis, testis, and lung showed a single RNA species at 1.3 kb is expressed in liver, intestine, and kidney. Guinea pig 3β-hydroxysteroid sulfotransferase is thus different from that in humans, who possess two mRNA species of 1.3 and 1.8
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