Defects in the conversion of androstenedione to testosterone in the fetal testes by the enzyme 17 beta-hydroxysteroid dehydrogenase (17 beta-HSD) give rise to genetic males with female external genitalia. We have used expression cloning to isolate cDNAs encoding a microsomal 17 beta-HSD type 3 isozyme that shares 23% sequence identity with other 17 beta-HSD enzymes, uses NADPh as a cofactor, and is expressed predominantly in the testes. The 17 beta HSD3 gene on chromosome 9q22 contains 11 exons. Four substitution and two splice junction mutations were identified in the 17 beta HSD3 genes of five unrelated male pseudohermaphrodites. The substitution mutations severely compromised the activity of the 17 beta-HSD type 3 isozyme.

Two isozymes of steroid 5a-reductase encoded by separate loci catalyze the conversion of testosterone to dihydrotestosterone. Inherited defects in the type 2 isozyme lead to male pseudohermaphroditism in which affected males have a normal internal urogenital tract but external genitalia resembling those of a female. The 5a-reductase type 2 gene (gene symbol SRD5A2) was cloned and shown to contain five exons and four introns. The gene was localized to chromosome 2 band p23 by somatic cell hybrid mapping and chromosomal in situ hybridization. Molecular analysis of the SRD5A2 gene resulted in the identification of 18 mutations in 11 homozygotes, 6 compound heterozygotes, and 4 inferred compound heterozygotes from 23 families with 5a-reductase deficiency. 6 apparent recurrent mutations were detected in 19 different ethnic backgrounds. In two patients, the catalytic efficiency of the mutant enzymes correlated with the severity of the disease. The high proportion of compound heterozygotes suggests that the carrier frequency of mutations in the 5a-reductase type 2 gene may be higher than previously thought. (J. Clin. Invest. 1992. 799-809.)

We describe a metabolic defect in bile acid synthesis involving a deficiency in 7 ␣ -hydroxylation due to a mutation in the gene for the microsomal oxysterol 7 ␣ -hydroxylase enzyme, active in the acidic pathway for bile acid synthesis. The defect, identified in a 10-wk-old boy presenting with severe cholestasis, cirrhosis, and liver synthetic failure, was established by fast atom bombardment ionization-mass spectrometry, which revealed elevated urinary bile acid excretion, a mass spectrum with intense ions at m/z 453 and m/z 510 corresponding to sulfate and glycosulfate conjugates of unsaturated monohydroxy-cholenoic acids, and an absence of primary bile acids. Gas chromatography-mass spectrometric analysis confirmed the major products of hepatic synthesis to be 3 ␤ -hydroxy-5-cholenoic and 3 ␤ -hydroxy-5-cholestenoic acids, which accounted for 96% of the total serum bile acids. Levels of 27-hydroxycholesterol were Ͼ 4,500 times normal. The biochemical findings were consistent with a deficiency in 7 ␣ -hydroxylation, leading to the accumulation of hepatotoxic unsaturated monohydroxy bile acids. Hepatic microsomal oxysterol 7 ␣ -hydroxylase activity was undetectable in the patient. Gene analysis revealed a cytosine to thymidine transition mutation in exon 5 that converts an arginine codon at position 388 to a stop codon.

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