LAUSR

Rare sugars and sugar derivatives that can be obtained from abundant sugars are of great interest to biochemical and pharmaceutical research. Here, we describe the substrate scope of a short‐chain dehydrogenase/reductase from Sphingomonas species A1 (SpsADH) in the oxidation of aldonates and polyols. The resulting products are rare uronic acids and rare sugars respectively. We provide insight into the substrate recognition of SpsADH using kinetic analyses, which show that the configuration of the hydroxyl groups adjacent to the oxidized carbon is crucial for substrate recognition. Furthermore, the specificity is demonstrated by the oxidation of d‐sorbitol leading to l‐gulose as sole product instead of a mixture of d‐glucose and l‐gulose. Finally, we applied the enzyme to the synthesis of l‐gulose from d‐sorbitol in an in vitro system using a NADH oxidase for cofactor recycling. This study shows the usefulness of exploring the substrate scope of enzymes to find new enzymatic reaction pathways from renewable resources to value‐added compounds.