Molecular Cloning, Nucleotide Sequence and Tissue Distribution of Equine Testicular 3.BETA.-Hydroxysteroid Dehydrogenase/.DELTA.5-.DELTA.4 Isomerase Messenger Ribonucleic Acid.

Abstract

Complementary DNA (cDNA) clones encoding equine testicular steroidogenic enzyme 3β-hydroxysteroid dehydrogenase/Δ5-Δ4 isomerase (3β-HSD) have been isolated from equine testicular cDNA library. Polymerase chain reaction (PCR) with 3β-HSD specific primers was performed with equine testicular cDNA, and PCR fragment was used as a probe for screening of equine testicular λ ZAP II phage cDNA library. Positive clones were excised into plasmid vector pBluescript and determined their nucleotide sequences. Total 1651 bp of the cDNA sequence was consisted with 112 bp of 5' flanking untranslated sequence, 1122 bp open reading frame encodes 373 amino acids, and putative poly-adenylation signal “AATAAA” at 385 bp downstream of the stop codon. The nucleotide and the deduced amino acid sequence of equine 3β-HSD were very similar to the known mammalian 3β-HSD sequences. From the deduced amino acid sequence, equine testicular 3β-HSD may be concluded to catalyze 3β-dehydrogenation and Δ-isomerization, but 3-ketoreduction as well as murine isoforms. As the clone was used in Northern blot analysis of equine tissues, the expression of equine 3β-HSD mRNA was prominent in the classical steroidogenic tissues including placenta.