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Updated: Jun 24, 2026

Characterization of Glycoproteins with the Immunoglobulin Fold by X-Ray Crystallography and Biophysical Techniques
Published on: July 5, 2018
Characterization of glycerol dehydratase expressed by fusing its alpha- and beta-subunits
Xu-De Wang1, Xian-En Zhang, Yong-Chao Guo
1State Key Laboratory of Virology, Wuhan Institute of Virology, Chinese Academy of Sciences, Wuhan, 430071, China.
Abstract:
The gdh and gdhr genes, encoding B(12)-dependent glycerol dehydratase (GDH) and glycerol dehydratase reactivase (GDHR), respectively, in Klebsiella pneumoniae, were cloned and expressed in E. coli. Part of the beta-subunit was lost during GDH purification when co-expressing alpha, beta and gamma subunit. This was overcome by fusing the beta-subunit to alpha- or gamma-subunit with/without the insertion of a linker peptide between the fusion moieties. The kinetic properties of the fusion enzymes were characterized and compared with wild type enzyme. The results demonstrated that the fusion protein GDHALB/C, constructed by linking the N-terminal of beta-subunit to the C-terminal of alpha subunit through a (Gly(4)Ser)(4) linker peptide, had the greatest catalytic activity. Similar to the wild-type enzyme, GDHALB/C underwent mechanism-based inactivation by glycerol during catalysis and could be reactivated by GDHR.
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