Abstract
Proof the formation of enzyme-substrate complex between DPN-specific dehydrogenases and coenzymes as well as their substrates were carried out by the method of “crossing paper electrophoresis”.
The enzymes used were lactic dehydrogenase (LDH), malic dehydrogenase (MDH), alcohol dehydrogenase (ADH), and glycerinaldehyde-3-phosphate dehydrogenase (GDH). On a filter paper, an enzyme solution was applied on a line drawn in parallel to the direction of the electrical field and a solution of substrate or coenzyme was applied on a line parpendicular to the former. After the electrophoresis the line of the substrate or coenzyme became hollow at the point of crossing with the line of enzyme. Thus the direct interaction and therefore the formation of complex between the enzyme and substrate as well as coenzymes were demonstrated with all the enzymes tested.
In regard to the substrates, lactic and pyruvic acids were applied to LDH, and malic acid to MDH, but to ADH and GDH no substrate was applied. On the other hand the preparation used of MDH was not pure, so that the proof in this case can not be regarded as complete.
