Isolation and characterization of the cDNA encoding the spiny dogfish shark ( Squalus acanthias) form of cytochrome P450c17
Abstract Cytochrome P450c17 is a key steroidogenic enzyme for the production of sex steroids in gonadal tissue and for cortisol production in adrenal tissue. This protein possesses two enzymatic activities. The 17α‐hydroxylase activity results in the introduction of a hydroxyl group at the 17α‐posit...
| Published in: | Journal of Experimental Zoology |
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| Main Author: | |
| Format: | Article in Journal/Newspaper |
| Language: | English |
| Published: |
Wiley
1995
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| Online Access: | http://dx.doi.org/10.1002/jez.1402720104 https://api.wiley.com/onlinelibrary/tdm/v1/articles/10.1002%2Fjez.1402720104 https://onlinelibrary.wiley.com/doi/pdf/10.1002/jez.1402720104 |
| Summary: | Abstract Cytochrome P450c17 is a key steroidogenic enzyme for the production of sex steroids in gonadal tissue and for cortisol production in adrenal tissue. This protein possesses two enzymatic activities. The 17α‐hydroxylase activity results in the introduction of a hydroxyl group at the 17α‐position. The resultant 17α‐hydroxylated, C21 pregnene is converted to a C19 andro‐gen by the C17,20‐lyase activity. A cDNA library was constructed from poly(A)‐enriched mRNA isolated from spiny dogfish shark ( Squalus acanthias ) testis and ligated into Eco RI‐cut lambda arms. The amplified library was screened using a bovine P450c17 cDNA probe and five positive clones were isolated. The described cDNA encompasses 23 bp of the 5′‐untranslated region, a 1,527 bp open reading frame, and 414 bp of the 3′‐untranslated region. A putative polyadenylation signal (AATAAA) is 18 bp from the poly(A) tail. Northern blot analysis showed a single transcript of 1.9 kb, thus indicating the isolated clone is a full‐length cDNA. The deduced amino acid sequence of the shark form of P450c17 is 59% and 57% identical to the rainbow trout and chicken forms, respectively. The shark form is 43% to 46% identical to mammalian forms (rat, human, mouse, bovine, and porcine). There are large regions of extremely high identity shared among all the forms. The deduced shark 17α‐hydroxylase protein is 509 residues in length with a predicted weight of 57.2 kDa. Non‐steroidogenic COS cells, transfected with the shark P450c17 cDNA, was capable of 17α‐hydroxylase and C17,20‐lyase activities using both pregnenolone and progesterone as initial substrates. © 1995 Wiley‐Liss, Inc. |
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