BIOTRANSFORMATION OF 13-CIS-RETINOIC AND 9-CIS-RETINOIC ACID TO ALL-TRANS-RETINOIC ACID IN RAT CONCEPTAL HOMOGENATES - EVIDENCE FOR CATALYSIS BY A CONCEPTAL ISOMERASE

The purpose of this study was to investigate whether and to what exten t the steric isomerization of retinoic acids in conceptal tissues can be attributed to enzymatic catalysis in addition to thioldependent, no nenzymatic catalysis. Conversions of 13-cis-retinoic acid and 9-cis-re tinoic acid to all-trans-retinoic acid catalyzed by cell-free preparat ions of conceptal rat tissues (gestational day 12.5) were investigated . Substrates and rat conceptal homogenates (RCH) were incubated in sod ium phosphate buffer (0.1 M, pH 7.5) at 37 degrees C in the dark. Incu bation mixtures were quantitatively analyzed by HPLC. In RCH-catalyzed reactions, conversions of 13-cis-retinoic acid or 9-cis-retinoic acid to all-trans-retinoic acid were very rapid, in comparison with uncata lyzed isomerization reactions (incubations without RCH). Comparisons o f the rates of reactions catalyzed by freshly prepared vs. freshly pre pared/dialyzed RCH showed no significant differences, indicating that small, suflhydryl-containing molecules such as reduced glutathione did not significantly contribute to the RCH-catalyzed reactions. Furtherm ore, at physiological concentrations (2.5-10 mM), reduced glutathione exhibited very low specific catalytic activities, indicating that none nzymatic, sulfhydryl-dependent catalysis was not a major mechanism in catalyzing Interconversions of retinoic acids in vivo. Enzymatic catal ysis by RCH of the conversion of 13-cis-retinoic acid to all-trans-ret inoic acid was further characterized by showing 1) substrate saturatio n kinetics, 2) reaction rates that increased proportionally with prote in concentrations, and (3) much greater sensitivity of the reactions t o heat inactivation and denaturation by urea, compared with nonenzymat ic, glutathione-catalyzed reactions. Thus, isomerization of retinoids in conceptal tissues appeared to be under enzymatic control.