Formation of a Covalent Adduct in Retaining β-Kdo Glycosyl-Transferase WbbB via Substrate-Mediated Proton Relay
Otros/as autores/as
Fecha de publicación
2024-07-22ISSN
1867-3899
Resumen
The GT99 domain of the membrane-anchored WbbB glycosyltransferase (WbbBGT99) catalyzes the transfer of 3-deoxy-D-manno-oct-2-acid (β-Kdo) to an O-antigen saccharide acceptor with retention of stereochemistry. It has been proposed that the enzyme follows an unprecedented double-displacement mechanism involving the formation of covalent adduct between the Kdo sugar and an active site residue (Asp232) that is properly oriented for nucleophilic attack. Here we use QM/MM metadynamics simulations on recently reported crystal structures to provide theoretical evidence for the formation of such adduct and unveil the atomic details of the chemical reaction. Our results support the interpretation made on the basis of X-ray and mass spectrometry analyses. Moreover, we show that the formation of the β-Kdo-Asp232 adduct is assisted by the sugar Kdo-carboxylate group, which mediates the transfer of a proton from Asp232 towards the phosphate leaving group, alleviating electrostatic repulsion between the two negatively charged carboxylate groups. The computed mechanism also explains why His265, previously proposed to act as a general acid, does not impair catalysis. This mechanism can be extended to other related enzymes, expanding the repertoire of GT mechanisms in Nature.
Tipo de documento
Artículo
Versión del documento
Versión publicada
Lengua
Inglés
Materias (CDU)
54 - Química
Palabras clave
3-deoxy-D-manno-oct-2-acid
carbohydrate-active enzymes
metadynamics
molecular dynamics
Quantum mechanics/molecular mechanics
Catalytic mechanism
Páginas
8 p.
Publicado por
Wiley
Publicado en
ChemCatChem. 2024;16:e202400769
Número del acuerdo de la subvención
info:eu-repo/grantAgreement/EC/ERC-2020-SyG-951231
info:eu-repo/grantAgreement/SUR del DEC i AGAUR/2021-SGR-00680
info:eu-repo/grantAgreement/MICIN, AEI i FEDER/PN I+D/PID2022-138252OB−I00
info:eu-repo/grantAgreement/MICIN, AEI i FEDER/PN I+D/PID2020-118893GB-100
info:eu-repo/grantAgreement/MICIN i AEI/María de Maeztu/CEX2021-001202-M
Este ítem aparece en la(s) siguiente(s) colección(ones)
Derechos
© L'autor/a
Excepto si se señala otra cosa, la licencia del ítem se describe como http://creativecommons.org/licenses/by-nc/4.0/