Employing the rabbit liver progesterone-21-hydroxylase P-450 1 cDNA as a probe (Tukey, R.H., Okino, S., Barnes, H., Griffin, K.J., and Johnson, E.F. (1985) J. Biol. Chem. 260, 13347-13354), we have identified a highly homologous (81% within the coding region) human liver cDNA, termed Hp1-1, that encodes a 490-amino acid protein. Comparison of the predicted translation products between the human and rabbit homologues demonstrates that the two proteins are 73% homologous, while increasing to 82% similarity when allowing for conserved amino changes. The human P-450 1 is 82% homologous to the s-mephenytoin 4-hydroxylase (Umbenhauer, D. R., Martin, M. V., Lloyd, R. S., and Guengerich, F. P. (1987) Biochemistry 26, 1094-1099). Southern blot analysis using various portions of the human P-450 1 cDNA as probes indicates that the human P-450 1 gene is part of a larger gene family but can be selectively identified by using a 3'-noncoding portion of the cDNA. Identification of the gene from a panel of human-rodent somatic cell hybrids using the conserved 3' portion of the human P-450 1 cDNA as a probe places the location of the gene on human chromosome 10. Results are also presented which demonstrate that the human P-450 1 gene transcript is processed by an alternate RNA-splicing mechanism that generates two mRNA products, one which represents the functional transcript, and the other a form of mRNA that is not capable of encoding a functional P-450.