Abstract
By sequence comparison, the majority of 1,4-α-glucan-branching enzymes (GBEs) consist of an N-terminal carbohydrate-binding domain, a TIM-barrel catalytic domain, and a C-terminal all-beta domain. Among these structures, the GBE from Geobacillus thermoglucosidans STB02 uniquely has a highly charged 26-amino-acid C-terminal extension, whose functional roles are the least understood. In this research, the functional significance of the C-terminal domain in GBE from G. thermoglucosidans STB02 and its extension were assessed using a C-terminal deletion analysis. Mutants lacking of more than 7 residues of the C-terminal all-beta domain could not be detected in lysates of their Escherichia coli expression strains, suggesting that an intact all-beta domain is required for structural stability. In contrast, truncation of the C-terminal extension resulted in greater stability and solubility than the wild type, as well as a lower sensitivity to the presence of added metal ions. Comparison of this mutant with the wild type suggests that the interaction of metal ions with the C-terminal extension influences performance of this enzyme.
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Funding
This was financially supported by the National Natural Science Foundation of China (No. 31722040, 31771935), the Natural Science Foundation of Jiangsu Province (BK20180606), and the China Postdoctoral Science Foundation (No. 2018M632233).
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Ban, X., Li, C., Zhang, Y. et al. Importance of C-Terminal Extension in Thermophilic 1,4-α-Glucan Branching Enzyme from Geobacillus thermoglucosidans STB02. Appl Biochem Biotechnol 190, 1010–1022 (2020). https://doi.org/10.1007/s12010-019-03150-7
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DOI: https://doi.org/10.1007/s12010-019-03150-7