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Publication Abstract from PubMed

Cellobiose 2-epimerase (CE) reversibly converts D-glucose residues into D-mannose residues at the reducing end of unmodified beta-1,4-linked oligosaccharides, including beta-1,4-mannobiose, cellobiose, and lactose. CE is responsible for conversion of beta-1,4-mannobiose to 4-O-beta-D-mannosyl-D-glucose in mannan metabolism. However, the detailed catalytic mechanism of CE is unclear due to the lack of structural data in complex with ligands. We determined the crystal structures of halothermophile Rhodothermus marinus CE (RmCE) in complex with substrates/products or intermediate analog, and its apo-form. The structures in complex with the substrates/products indicated that the residues in the beta5-beta6 loop as well as those in the inner six helixes form the catalytic site. Trp322 and Trp385 interact with reducing and non-reducing end parts of these ligands, respectively, by stacking interactions. The architecture of the catalytic site also provided insights into the mechanism of reversible epimerization. His259 abstracts the H2 proton of the D-mannose residue at the reducing end, and consistently forms the cis-enediol intermediate by facilitated depolarization of the 2-OH group mediated by hydrogen bonding interaction with His200. His390 subsequently donates the proton to the C2 atom of the intermediate to form a D-glucose residue. The reverse reaction is mediated by these three histidines with the inverse roles of acid/base catalysts. The conformation of cellobiitol demonstrated that the deprotonation/reprotonation step is coupled with rotation of the C2-C3 bond of the open-form of the ligand. Moreover, it is postulated that His390 is closely related to ring opening/closure by transferring a proton between the O5 and O1 atoms of the ligand.

Structural insights into the epimerization of beta-1,4-linked oligosaccharides catalyzed by cellobiose 2-epimerase, the sole enzyme epimerizing non-anomeric hydroxyl groups of unmodified sugars.,Fujiwara T, Saburi W, Matsui H, Mori H, Yao M J Biol Chem. 2013 Dec 20. PMID:24362032^[1]

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.