Cofactor recycling in a coupled enzyme oxidation-reduction reaction: conversion of omega-oxo-fatty acids into omega-hydroxy and dicarboxylic acids

Aldehydes are reduced to alcohols by the enzyme alcohol dehydrogenase (ADH) , whereas the enzyme aldehyde dehydrogenase (AldDH) oxidizes aldehydes to c arboxylic acids. ADH and AldDH require, respectively, the reduced and oxidi zed forms of the cofactor NAD (NAD(+)/NADH). By combining both oxidation an d reduction reactions into one process, it is possible to produce alcohols and carboxylic acids simultaneously from aldehydes by continuous recycling of the NAD(+)/ NADH cofactor. However, both enzymes need to be active withi n the same pH region and buffer system. To test this hypothesis, the pH pro file (Y-max and V-max/K-m) as well as the pK(a) of the prototropic groups i nvolved in catalysis for both dehydrogenases were determined using (Z,Z)-no na-2,4-dienal as a model substrate. The pH profile (V-max and V-max/K-m) of both enzymes overlapped in the pH range of 6-8 in potassium phosphate buff er. When the coupled enzyme system was used at pH 7 with 10% NAD(+) cofacto r, over 90% of the starting aldehyde was converted to its corresponding aci d and alcohol derivatives in a I : I ratio, The sequential action of the en zymes lipoxygenase and hydroperoxide lyase converts polyunsaturated fatty a cids to aldehydic fatty acids. The products arising from the oxidation or r eduction of the aldehydic functionality are of industrial interest. It was found that 13-oxo-9-(Z),11-(E)-tridecadienoic acid, the product of the sequ ential reaction of soya bean lipoxygenase and hydroperoxide lyase from Chlo rella pyrenoidosa on linoleic acid, is also a substrate in this coupled enz yme system.