Collect. Czech. Chem. Commun.
1986, 51, 1542-1549
https://doi.org/10.1135/cccc19861542
The use of Spheron as a matrix for affinity chromatography of NAD-dependent dehydrogenases
Jan Kovář, Alena Škodová and Jaroslav Turánek
Department of Biochemistry, Purkyně University, 611 37 Brno
Abstract
The paper compares several methods of coupling common ligands of dehydrogenases, viz. N6-[(6-aminohexyl)carbamoylmethyl]-AMP and N6-[(6-aminohexyl)carbamoylmethyl]-NAD, to a hydrophilic macroporous glycolmethacrylate gel, Spheron. The affinants coupled best to a CNBr-activated gel and to a gel with hydrazine groups (after activation with nitrous acid). The affinity properties of gels based on Spheron and on Sepharose 4B were similar ( the stability and separation efficiency were almost identical, the binding capacity and the recovery of dehydrogenase activity were somewhat better with the Sepharose). The materials based on Spheron were used in several separation experiments, viz. separation of lactate dehydrogenase form albumin, separation of lactate dehydrogenase from alcohol dehydrogenase under different conditions and separation of isoenzymes of lactate dehydrogenase. Spheron 300 with a coupled affinant was also employed in an attempt to purify a crude alcohol dehydrogenase.
