Michael Waterman
Faculty Member
Last active: 2/12/2015

Characterization of complementary deoxyribonucleic acid for human adrenocortical 17 alpha-hydroxylase: a probe for analysis of 17 alpha-hydroxylase deficiency.

Bradshaw KD, Waterman MR, Couch RT, Simpson ER, Zuber MX
Mol Endocrinol. 1987 1 (5): 348-54

PMID: 3274893 · DOI:10.1210/mend-1-5-348

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.

MeSH Terms (16)

Adrenal Hyperplasia, Congenital Amino Acid Sequence Animals Base Sequence Cell Line Cloning, Molecular DNA Humans Molecular Sequence Data Restriction Mapping RNA, Messenger Sequence Homology, Nucleic Acid Skin Steroid 17-alpha-Hydroxylase Steroid Hydroxylases Transfection

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