9RL0

CDP-tyvelose 2-epimerase from Thermodesulfatator atlanticus

Summary for 9RL0

• Entry DOI: 10.2210/pdb9rl0/pdb
• Descriptor: CDP-tyvelose 2-epimerase, CYTIDINE-5'-DIPHOSPHATE, NICOTINAMIDE-ADENINE-DINUCLEOTIDE, ... (4 entities in total)
• Functional Keywords: cdp-tyvelose 2-epimerase, sugar binding protein
• Biological source: Thermodesulfatator atlanticus
• Total number of polymer chains: 8
• Total formula weight: 321428.28

Authors

Rapp, C.,van Overtveldt, S.,Pfeiffer, M.,Beerens, K.,Merkas, M.,Pavkov-Keller, T.,Desmet, T.,Nidetzky, B. (deposition date: 2025-06-16, release date: 2026-06-10)

Primary citation

Rapp, C.,van Overtveldt, S.,Sanchez-Murcia, P.A.,Pfeiffer, M.,Beerens, K.,Merkas, M.,Pavkov-Keller, T.,Desmet, T.,Nidetzky, B.
A dual catalytic architecture promotes C-2 stereoinversion of CDP-glucose by the CDP-tyvelose 2-epimerase from Thermodesulfatator atlanticus.
J.Biol.Chem., 302:111384-111384, 2026

PubMed Abstract: The CDP-tyvelose 2-epimerase from Thermodesulfatator atlanticus (TaTyvE) catalyzes the C-2 epimerization of CDP-glucose to CDP-mannose. The enzyme uses NAD-dependent oxidation-reduction to achieve C-2 configurational inversion of the substrate. Here, we report the 2.60-Å crystal structure of tetrameric TaTyvE with NAD bound in all subunits and CDP bound in one (Protein Data Bank code: 9RL0). Binding of CDP orders the Gly197-Trp207 loop, closing over the sugar binding pocket. The active site of the ternary complex is well preorganized, with only moderate induced-fit conformational changes. Molecular dynamics simulations and site-directed mutagenesis suggest that TaTyvE employs a dual catalytic architecture to control substrate specificity. Asn125, within the TNK segment (Thr124-Asn125-Lys126), promotes sampling of catalytically plausible glucose conformations. The VAM segment (Val83-Ala84-Met85) permits broader conformational flexibility for mannose through backbone contacts. This is supported by a modest ∼12-fold activity reduction in the Q205A variant. In contrast, N125A abolishes activity. Substrate analogs featuring deoxygenation, stereoinversion, or fluorination at the C-4 were synthesized to examine the role of the sugar C4-OH. Simulations indicated that the C4-OH interacts with Val83, Asn125, and Gln205. Activity was lost with CDP-4-deoxy-glucose and minimally recovered with CDP-4-fluoro-glucose. Removing the C5-hydroxymethyl group to restrict substrate positioning flexibility enhanced the reaction rate. Overall, these results highlight the important interplay of structural preorganization and conformational flexibility in TaTyvE for enzyme activity and specificity in the C-2 epimerization of CDP-glucose.

PubMed: 41861995
DOI: 10.1016/j.jbc.2026.111384

Experimental method

X-RAY DIFFRACTION (2.6 Å)