The Conversion of UDP‐Glc to UDP‐Man: In Silico and Biochemical Exploration To Improve the Catalytic Efficiency of CDP‐Tyvelose C2‐Epimerases

Abstract

A promiscuous CDP‐tyvelose 2‐epimerase (TyvE) from Thermodesulfatator atlanticus (TaTyvE) belonging to the Nucleotide Sugar active Short‐Chain Dehydrogenase/Reductase superfamily (NS‐SDRs) was recently discovered. TaTyvE performs the slow conversion of NDP‐glucose (NDP‐Glc) to NDP‐mannose (NDP‐Man). Here, we present the sequence fingerprints that are indicative of the conversion of UDP‐Glc to UDP‐Man in TyvE‐like enzymes based on the heptagonal box motifs. Our data‐mining approach led to the identification of 11 additional TyvE‐like enzymes for the conversion of UDP‐Glc to UDP‐Man. We characterized the top two wild‐type candidates that show a 15‐ and 20‐fold improved catalytic efficiency on UDP‐Glc compared to TaTyvE. Additionally, we present a quadruple variant of one of the identified enzymes with a 70‐fold improved catalytic efficiency on UDP‐Glc compared to TaTyvE. These findings may help the design of new nucleotide production pathways starting from a cheap sugar substrate like glucose or sucrose.