Coenzyme-dependent enzymatic synthesis of fine chemicals. Enzymatic synthesis of L-ascorbic acid via uronic acids. 2. L-hexonate dehydrogenase from yeasts

Abstract

Enzymatic synthesis of L-ascorbic acid may be carried out starting from uronic acids by applying a system with intrase quential cofactor regeneration (figure 2). The first step in this pathway to ascorbic acid is the reduction of D-glucuronic acid to L-gulonic acid by L-hexonate dehydrogenase. This enzyme was detected in the cytoplasma of several yeasts (Lipomyces starkeyi, Saccharomyces cerevisiae, Schwanniomyces occidentalis and Saccharomyces lipolytica). Up to date all the enzymes found are NADPH-dependent. The enzyme activities varied considerably with media composition and time of harvest. Until now, the best results were obtained for L. starkeyi grown on glucose, galactose or myo-inositol. The enzyme of L. starkeyi was purified by hydrophobic interaction chromatography on Phenyl-Sepharose CL 4B and anion-exchange chromatography (FPLC: Mono Q). The protein has a pH-optimum around pH 7 and a molecular weight of 30.000 D (as determined on a Superose 12 FPLC-column). At present, the enzyme is characterized in more detail