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Role of N-linked glycosylation in the enzymatic properties of a thermophilic GH 10 xylanase from Aspergillus fumigatus expressed in Pichia pastoris.


ABSTRACT: N-Glycosylation is a posttranslational modification commonly occurred in fungi and plays roles in a variety of enzyme functions. In this study, a xylanase (Af-XYNA) of glycoside hydrolase (GH) family 10 from Aspergillus fumigatus harboring three potential N-glycosylation sites (N87, N124 and N335) was heterologously produced in Pichia pastoris. The N-glycosylated Af-XYNA (WT) exhibited favorable temperature and pH optima (75°C and pH 5.0) and good thermostability (maintaining stable at 60°C). To reveal the role of N-glycosylation on Af-XYNA, the enzyme was deglycosylated by endo-?-N-acetylglucosaminidase H (DE) or modified by site-directed mutagenesis at N124 (N124T). The deglycosylated DE and mutant N124T showed narrower pH adaptation range, lower specific activity, and worse pH and thermal stability. Further thermodynamic analysis revealed that the enzyme with higher N-glycosylation degree was more thermostable. This study demonstrated that the effects of glycosylation at different degrees and sites were diverse, in which the glycan linked to N124 played a key role in pH and thermal stability of Af-XYNA.

SUBMITTER: Chang X 

PROVIDER: S-EPMC5302446 | biostudies-literature | 2017

REPOSITORIES: biostudies-literature

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Role of N-linked glycosylation in the enzymatic properties of a thermophilic GH 10 xylanase from Aspergillus fumigatus expressed in Pichia pastoris.

Chang Xiaoyu X   Xu Bo B   Bai Yingguo Y   Luo Huiying H   Ma Rui R   Shi Pengjun P   Yao Bin B  

PloS one 20170210 2


N-Glycosylation is a posttranslational modification commonly occurred in fungi and plays roles in a variety of enzyme functions. In this study, a xylanase (Af-XYNA) of glycoside hydrolase (GH) family 10 from Aspergillus fumigatus harboring three potential N-glycosylation sites (N87, N124 and N335) was heterologously produced in Pichia pastoris. The N-glycosylated Af-XYNA (WT) exhibited favorable temperature and pH optima (75°C and pH 5.0) and good thermostability (maintaining stable at 60°C). To  ...[more]

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