Feb 22, 1988 - ... sulfates at the 1513- position and the male-specific 16a-hydroxylase (P-450 16a) .... at 65°C sequentially with 2 x SSC/0.5 x SSC/0.1 x SSC.
Proc. Nati. Acad. Sci. USA
Vol. 85, pp. 4214-4217, June 1988 Biochemistry
cDNA cloning, sequence, and regulation of a major female-specific and growth hormone-inducible rat liver cytochrome P-450 active in 15 f-hydroxylation of steroid sulfates (library screening/genomic hybridization/homology of P450 cytochromes/sexually dimorphic growth hormone pattern)
PETER G. ZAPHIROPOULOS, AGNETA MODE, ANDERS STR6M, CHRISTER MOLLER, CARMEN FERNANDEZ, AND JAN-AKE GUSTAFSSON Department of Medical Nutrition, F69, and Center for BioTechnology, Huddinge University Hospital, S-141 86 Huddinge, Sweden
Communicated by Viktor Mutt, February 22, 1988
found to be dependent on the plasma pattern of growth hormone (GH) (6, 7). GH is secreted in a sex-dependent manner; in males, high GH surges occur at 3- to 4-hr intervals with low, often undetectable, levels in between, whereas in females there is an almost continuous presence of GH in serum due to more frequent and irregular peaks (8). By mimicking the male- and female-specific secretory pattern by intermittent or continuous administration of GH to hypophysectomized animals, a differential expression of P-450 1513 and P-450 16a has been achieved. Recently, cDNA clones for P-450 151, (9) and P-450 16a (10) have been isolated in this laboratory and used to quantitate respective mRNA levels after different GH treatments. The results indicated that GH regulation of these two cytochromes occurs at a pretranslational level. In this report, we describe the isolation and sequencing of cDNA clones encompassing the complete coding sequence of cytochrome P-450 1513, the structural relationships of this female-specific enzyme with the male-specific cytochrome P450 16a as well as with other members of the P-450 gene superfamily, and the time course of induction of these two sex-specific cytochromes by the secretory pattern of GH.t
cDNA clones encompassing the complete codABSTRACT ing sequence for the female-specific rat liver microsomal 15.8-hydroxylase have been isolated and sequenced. This cytochrome P-450 species (P.450 15/3) has a total of 490 amino acid residues, and its deduced amino acid sequence as well as direct sequencing of the amino-terminal portion of the purified protein revealed its identity with P450 i. P-450 15/3 was found to be 66% similar to the male-specific rat liver microsomal 16a-hydroxylase, P-450 16a. Furthermore, P-450 15.8 is 47% similar to the major phenobarbital-inducible cytochrome P450 in rat liver, P-450 b, while its structural similarity to P450 c, P450 pregnenolone-16a-carbonitrile, and P450 lauric acid w-hydroxylase is
