Many research areas, e.g., basic research but also applied fields of biotechnology, biomedicine, and
diagnostics often suffer from the unavailability of metabolic compounds. This is mostly due to missing
easy and efficient synthesis procedures. We herein describe the biocatalytic/enzymatic production of
2-keto-3-deoxy-D-gluconate, an intermediate of central metabolic pathways in all three domains of life
and also of bacterial polysaccharides, lipopolysaccharides, and cell wall components. The method is based
on the gluconate dehydratase from the hyperthermophilic crenarchaeon Thermoproteus tenax, which can be
easily recombinantly overproduced in Escherichia coli and—due to its intrinsic thermostability—rapidly be
purified by two precipitation steps. The enzyme completely converts D-gluconate to solely stereochemically
pure KDG, taking benefits from the enol-keto-tautomerism of the primary reaction product. The final
product can then easily be separated from the protein by ultrafiltration. The simple one-step procedure,
which is suitable at least for the lab-scale/gram-scale production of KDG, replaces lengthy multi-step
reactions and is easily scalable. This approach also illustrates the great application potential of Archaea with
their unusual metabolic pathways and enzymes for the synthesis of added value products.
Höfmann, S. et al. (2022). Simplified Enzymatic Synthesis of 2-Keto-3-Deoxy-D-Gluconate from D-Gluconate Using the Gluconate Dehydratase from Thermoproteus tenax. In: Ferreira-Cerca, S. (eds) Archaea. Methods in Molecular Biology, vol 2522. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-2445-6_23
Link to chapter: https://link.springer.com/protocol/10.1007/978-1-0716-2445-6_23