A two-step purification of cytochrome P-450 from adult pig testis by pregnenolone affinity column chromatography

Adult testicular cytochrome P-450 was purified by a two-step procedure util izing hydroxylapatite and pregnenolone affinity column chromatography. The cytochrome P-450 was determined to have an isoelectric point of 6.43 on ana lytical isoelectric focusing. The purified cytochrome P-450 was found to be homogeneous and its molecular weight was estimated to be 52,000 on polyacr ylamide gel electrophoresis in the presence of sodium dodecyl sulfate. The carbon monoxide difference spectrum with a peak at 448 nm exhibited the abs orption spectrum of a typical cytochrome p-450. A purification of 755 x was achieved with a yield of 3.09%. Copyright (C) 1999 John Wiley & Sons, Ltd.