Kinetics of collagen‐bound sorbitol dehydrogenase in the rotating membrane reactor: Opposite variations of affinity constants under diffusional limitations

Abstract

Abstract Sorbitol dehydrogenase was bound to the surface of acyl‐azide‐activated collagen membranes and its kinetics was investigated as a model of two‐substrate or cofactor‐requiring enzyme reactions. The study was performed with the “rotating membrane reactor” especially designed to obtain a precise variation of the external mass‐transfer coefficient, and thus the direct visualization of diffusional effects on the bound enzyme behavior. Diffusional limitations for NADH were found to decrease the apparent affinity for NADH, but to increase the apparent affinity for fructose. Such opposite effects of diffusional limitations on apparent affinities are generally applicable to reactions involving two substrates or a substrate and a cofactor of widely different affinities.