"Affinity" chromatography of steroid-transforming enzymes with a non-steroidal ligand.
The chromatographic behaviour of an avian oestradiol-17 beta dehydrogenase, the 3(17) beta-hydroxy steroid dehydrogenase from Pseudomonas testosteroni and cortisone reductase from Streptomyces dehydrogenans was studied on columns of p-(phenoxypropoxy)aniline attached to CNBr-activated Sepharose. The ligand was effective in adsorbing the oestradiol dehydrogenase from a partially purified extract of chicken liver, and the cortisone reductase was perferentially retained when mixtures of the three dehydrogenases were applied to columns in 10mM-buffer. Under these conditions the 3(17)beta-hydroxy steroid dehydrogenase was eluted in the front, but was adsorbed in the presence of 3 M-KCl. beta-N-Acetylglucosaminidase present in the liver preparation was not retained by the ligand, whereas lactate dehydrogenase from rabbit muscle was adsorbed in a manner similar to the retention pattern found on affinity chromatography with 2',5'-ADP--Sepharose. The mean overall purification of the oestradiol dehydrogenase was 13-fold, with a mean recovery of 53%. p-(Phenoxypropoxy)aniline offers promise for the purification of steroid-transforming enzymes where elution with substrate or cofactor is not wanted. It is also suggested that the ligand may be of service in the purification of receptors of hormonal steroids.
D015067:17-Hydroxysteroid Dehydrogenases / Q000302:isolation & purification*; D015089:20-Hydroxysteroid Dehydrogenases / Q000302:isolation & purification*; D000118:Acetylglucosaminidase / Q000302:isolation & purification; D000814:Aniline Compounds; D002846:Chromatography, Affinity; D003349:Cortisone Reductase / Q000302:isolation & purification*; D004960:Estradiol Dehydrogenases / Q000302:isolation & purification*; D007770:L-Lactate Dehydrogenase / Q000302:isolation & purification; D008024:Ligands
D000814:Aniline Compounds; D008024:Ligands; C019042:4-(phenoxypropoxy)aniline; D015067:17-Hydroxysteroid Dehydrogenases; D015089:20-Hydroxysteroid Dehydrogenases; D007770:L-Lactate Dehydrogenase; D003349:Cortisone Reductase; D004960:Estradiol Dehydrogenases; D000118:Acetylglucosaminidase