RGD Reference Report - Molecular cloning of cDNA for rat ovarian 20 alpha-hydroxysteroid dehydrogenase (HSD1). - Rat Genome Database

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Molecular cloning of cDNA for rat ovarian 20 alpha-hydroxysteroid dehydrogenase (HSD1).

Authors: Miura, R  Shiota, K  Noda, K  Yagi, S  Ogawa, T  Takahashi, M 
Citation: Miura R, etal., Biochem J 1994 Apr 15;299 ( Pt 2):561-7.
RGD ID: 1299435
Pubmed: PMID:8172618   (View Abstract at PubMed)
PMCID: PMC1138308   (View Article at PubMed Central)

20 alpha-Hydroxysteroid dehydrogenase (20 alpha-HSD, EC 1.1.1.149) catalyses the conversion of progesterone into 20 alpha-dihydroprogesterone (20 alpha-OHP). Previously, we purified the enzyme (37 kDa) from rat ovary and determined its N-terminal amino acid sequence. In the present study we succeeded in cloning a full-length 20 alpha-HSD cDNA. mRNA was extracted from immature rat ovaries after successive treatment with equine chorionic gonadotropin (eCG) and human chorionic gonadotropin (hCG). A cDNA library was constructed in lambda ZAP. For screening, a 576 bp probe was amplified by the PCR using mixed primers based on the N-terminal sequence of 20 alpha-HSD, and labelled with [32P]dCTP. Eight positive clones were isolated from 1.2 x 10(4) recombinants. Analysis of the nucleotide sequence revealed that one clone of 1.2 kbp cDNA (pHSD12-07) contained a polyadenylation site and an open reading frame encoding 323 amino acids with the N-terminal sequence of 20 alpha-HSD. The fusion protein of pHSD12-07 produced by Escherichia coli reacted with a specific polyclonal antibody generated against rat ovarian 20 alpha-HSD. In addition, the in vitro transcription-translation product produced by Xenopus oocytes showed 20 alpha-HSD activity and Northern-blotting analysis revealed that the ovaries from normal adult rats contained a 1.2 kb mRNA. Thus we succeeded in isolating a clone encoding the full length of rat ovarian 20 alpha-HSD. The sequence showed high similarity with those of rat liver 3 alpha-hydroxysteroid dehydrogenase (3 alpha-HSD), bovine lung prostaglandin F synthase (PGFS), human liver chlordecone reductase (CDR), frog lens rho-crystallin and aldose reductases, indicating that 20 alpha-HSD belongs to the aldo-keto reductase family.

Gene Ontology Annotations    Click to see Annotation Detail View

Biological Process
TermQualifierEvidenceWithReferenceNotesSourceOriginal Reference(s)
progesterone metabolic process acts_upstream_of_or_withinIDA 1299435PMID:8172618MGI 

Molecular Function

Objects Annotated

Genes (Rattus norvegicus)
Akr1c3  (aldo-keto reductase family 1, member C3)


Additional Information