One-step synthesis of 2-keto-3-deoxy-d-gluconate by biocatalytic dehydration of d-gluconate

Abstract

2-Keto-3-deoxy-sugar acids are key intermediates of central metabolism and integral constituents of bacterial (lipo)polysaccharides and cell wall components and are therefore continuously and highly demanded in related research fields. The stereospecific chemical synthesis of chiral 2-keto-deoxy-sugar acids involves a multitude of reaction steps, while in metabolic pathways only few conversions lead to the same 2-keto-3-deoxy sugar acids from easily available carbohydrate precursors. Here we present a straightforward and highly economic one-step biocatalytic synthesis procedure of 2-keto-3-deoxy-d-gluconate (KDG) from d-gluconate using recombinant gluconate dehydratase (GAD) from the hyperthermophilic crenarchaeon Thermoproteus tenax. This method is highly advantageous to KDG production schemes described so far for several reasons: (i) the d-gluconate is completely converted to stereochemically pure D-KDG without side-product formation, (ii) the final KDG yield is approximately 90%, (iii) the newly developed quantitative and qualitative LC-MS analysis method enabled the simultaneous detection of d-gluconate and KDG and (iv) the T. tenax GAD as biocatalyst can be provided by a simple and rapid procedure involving only two precipitation steps. The described utilization of dehydratases for 2-keto-3-deoxy sugar acid syntheses represents a highly resource-efficient one-step preparation and offers potential short synthetic routes toward a broad range of 2-keto-3-deoxy sugar acids and their derivatives.

Keywords: 2-Keto-3-deoxy-d-gluconate; Biocatalytic dehydration; Gluconate dehydratase; LC–MS; Metabolite.