Preparation of a NAD(H)‐polymer matrix showing coenzymic function of the bound pyridine nucleotide

Abstract

Abstract To a Sepharose gel the pyridine nucleotide NAD(H) has been bound using dicyclohexyl carbodiimide. In order to improve the steric availability of the nucleotide for added soluble enzymes such as dehydrogenases, a spacer molecule, ε‐amino caproic acid, was inserted between the carbohydrate matrix and the nucleotide. The obtained preparation contained 56 μmoles NAD + /g dry polymer. The obtained matrix‐bound NAD(H) was accepted as coenzyme by added lactate dehydrogenase. These preparations were still active after storage for several weeks at 4° C and could be used repeatedly without loss of activity. This represents the first necessary step taken in the preparation of compact closed systems consisting of “enzyme–coenzyme–coenzyme‐regenerating enzyme” bound to individual polymer beads; such systems eliminate the need for continuous coenzyme addition.