Wataru Saburi, Hirohiko Muto-Fukiya, Nongluck Jaito, Koji Kato, Jian Yu, Min Yao, Haruhide Mori
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引用次数: 0
Abstract
Cellobiose 2-epimerase (CE) catalyzes C-2 epimerization of reducing end d-glucose/d-mannose residue of β-(1→4)-disaccharides, and also slightly catalyzes aldose-ketose conversion. In this study, we investigated the structure-function relationship of Rhodothermus marinus CE (RmCEs). In 2H2O, 2H replaced the 2-H of the reducing end sugar residue, suggesting a proton abstraction-addition mechanism via the cis-enediolate intermediate. The structure of the RmCE-mannobiitol complex showed that His259 was suitable for abstracting 2-H from d-mannose residue, whereas His390 was suitable for the d-glucose residue. H259A and H390A mutations abolished activity for Galβ1-4Man and Galβ1-4Glc formation from Galβ1-4Fru, respectively, and these mutants catalyzed both epimerization and isomerization to Galβ1-4Glc and Galβ1-4Man, respectively. Ala substitution of the residues interacting with the 2-O of the reducing end sugar residue significantly reduced the velocity for epimerization, but not for isomerization. Trp385, stacked onto the non-reducing-end sugar residues of disaccharides, was shown to be important for disaccharide specificity.
期刊介绍:
Bioscience, Biotechnology, and Biochemistry publishes high-quality papers providing chemical and biological analyses of vital phenomena exhibited by animals, plants, and microorganisms, the chemical structures and functions of their products, and related matters. The Journal plays a major role in communicating to a global audience outstanding basic and applied research in all fields subsumed by the Japan Society for Bioscience, Biotechnology, and Agrochemistry (JSBBA).