title: Characterization of complementary deoxyribonucleic acid for human adrenocortical 17 alpha-hydroxylase: a probe for analysis of 17 alpha-hydroxylase deficiency.
abstract: To provide a basis for investigation of the molecular mechanisms underlying the hormonal regulation of steroid 17 alpha-hydroxylase (P-450 17 alpha) activity in adrenal, ovary, and testis as well as human 17 alpha-hydroxylase deficiency, we have isolated from a human fetal adrenal cDNA library a cDNA sequence complementary to the mRNA that encodes the human P-450 17 alpha enzyme. Of 75,000 colonies from the library that were screened by use of a nick-translated 5'-specific bovine P-450 17 alpha cDNA probe, 10 positive colonies were isolated and the clone with the longest insert (pcD-17 alpha H) was selected for further characterization. pcD-17 alpha H encodes the complete human P-450 17 alpha protein having approximately 78% homology at the nucleotide level and 71% homology at the amino acid level when the sequence of pcD-17 alpha H is compared to the bovine P-450 17 alpha cDNA sequence. By transient expression of the human P-450 17 alpha cDNA clone in COS 1 cells, we have demonstrated that the 17 alpha-hydroxylase and 17,20 lyase activities reside within the same human P-450 17 alpha polypeptide chain. The insert was also used as a probe to investigate, by means of Southern blot analysis, possible alterations in the P-450 17 alpha gene sequence in DNA isolated from skin fibroblasts from three patients with clinically characterized 17 alpha-hydroxylase deficiencies. No changes were detected in the DNA of any of the patients by this analysis.
authors.affiliation: Cecil H. and Ida Green Center for Reproductive Biology Sciences, University of Texas Health Science Center, Dallas 75235.
authors.forename: K D
authors.initials: KD
authors.lastname: Bradshaw
authors.forename: M R
authors.initials: MR
authors.lastname: Waterman
authors.forename: R T
authors.initials: RT
authors.lastname: Couch
authors.forename: E R
authors.initials: ER
authors.lastname: Simpson
authors.forename: M X
authors.initials: MX
authors.lastname: Zuber
mesh_terms: D000312:Adrenal Hyperplasia, Congenital; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D003001:Cloning, Molecular; D004247:DNA / Q000235:genetics*; D006801:Humans; D008969:Molecular Sequence Data; D012333:RNA, Messenger / Q000235:genetics; D015183:Restriction Mapping; D012689:Sequence Homology, Nucleic Acid; D012867:Skin / Q000201:enzymology; D013254:Steroid 17-alpha-Hydroxylase / Q000235:genetics*; D013250:Steroid Hydroxylases / Q000235:genetics*; D014162:Transfection
chemical_list: D012333:RNA, Messenger; D004247:DNA; D013250:Steroid Hydroxylases; D013254:Steroid 17-alpha-Hydroxylase