Kinetic studies on the reactivation of D-beta-hydroxybutyrate dehydrogenase with mixtures of short-chain lecithins.
D-beta-hydroxybutyrate dehydrogenase, purified as soluble, lipid-free apoenzyme (inactive) from rat liver mitochondria can be reactivated by the short-chain dihexanoyl, diheptanoyl, and dioctanoyl lecithins at the monomeric state, upon formation of a reversible enzyme-lecithin complex. Previous studies with these lecithins suggested that reactivation of the apoenzyme requires the simultaneous occupation of two identical, noninteracting lecithin binding sites via a rapid equilibrium random mechanism. The short-chain lecithins exhibited similar reactivating capacities, differing only in their affinities towards the enzyme. In order to further test that model, the reactivation of the apoenzyme was studied when two or three short-chain lecithins were simultaneously present in the reaction medium. The initial velocities were measured either as a function of the concentration of one lecithin while the other(s) were kept constant, or as a function of the total phospholipid concentration with mixtures of different lecithins at a constant molar ratio. The pertinent equations were derived on the principles of multiple equilibria with identical, noninteracting sites able to be occupied by any of the different lecithins present in the reaction medium, with the doubly occupied enzyme as the only active species. In agreement with the above-proposed model, the results obtained indicates that the molar fraction of the doubly occupied (active) enzyme species can be calculated from equilibrium considerations and that the maximal attainable with the different short-chain lecithins are similar.
D000818:Animals; D055598:Chemical Phenomena; D002621:Chemistry; D004793:Enzyme Reactivators / Q000494:pharmacology*; D006884:Hydroxybutyrate Dehydrogenase / Q000378:metabolism*; D007700:Kinetics; D008433:Mathematics; D008930:Mitochondria, Liver / Q000201:enzymology; D008956:Models, Chemical; D010713:Phosphatidylcholines / Q000494:pharmacology*; D051381:Rats
D004793:Enzyme Reactivators; D010713:Phosphatidylcholines; D006884:Hydroxybutyrate Dehydrogenase