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N-glycosylation affects the proper folding, enzymatic characteristics and production of a fungal [beta]-glucosidase

Byline: Wei Wei, Ling Chen, Gen Zou, Qianfu Wang, Xing Yan, Jun Zhang, Chengshu Wang, Zhihua Zhou ABSTRACT Heterologous expression of [beta]-glucosidase is one of the approaches to enhance the efficiency of fungal cellulase preparations. It has been reported that N-glycosylation affects the structur... Full description

Journal Title: Biotechnology and Bioengineering Dec, 2013, Vol.110(12), p.3075(10)
Main Author: Wei, Wei
Other Authors: Chen, Ling , Zou, Gen , Wang, Qianfu , Yan, Xing , Zhang, Jun , Wang, Chengshu , Zhou, Zhihua
Format: Electronic Article Electronic Article
Language: English
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ID: ISSN: 0006-3592
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recordid: gale_ofa349975116
title: N-glycosylation affects the proper folding, enzymatic characteristics and production of a fungal [beta]-glucosidase
format: Article
creator:
  • Wei, Wei
  • Chen, Ling
  • Zou, Gen
  • Wang, Qianfu
  • Yan, Xing
  • Zhang, Jun
  • Wang, Chengshu
  • Zhou, Zhihua
subjects:
  • Monosaccharides
  • Enzymes
  • Asparagine
ispartof: Biotechnology and Bioengineering, Dec, 2013, Vol.110(12), p.3075(10)
description: Byline: Wei Wei, Ling Chen, Gen Zou, Qianfu Wang, Xing Yan, Jun Zhang, Chengshu Wang, Zhihua Zhou ABSTRACT Heterologous expression of [beta]-glucosidase is one of the approaches to enhance the efficiency of fungal cellulase preparations. It has been reported that N-glycosylation affects the structure framework, function and stability of proteins. In this study, a [beta]-glucosidase from Aspergillus terreus (GenBank: XP_001216552, BglS) was heterologously expressed in Pichia pastoris and Trichoderma reesei. The four asparagine residues were all linked with high-mannose-type oligosaccharides in P. pastoris, whereas only N224 carried high-mannose-type glycan in T. reesei (the other three sites carried one N-acetylglucosamine). The long N-glycan chains on PpBglS weakened its substrate affinity, activity and thermostability. The moderate post-translational and post-secretory glycan modification in T. reesei makes it a suitable expression system for BglS. The N224 glycan played a critical role in BglS folding. The elucidation of the correlation between the different N-glycosylation patterns of BglS and their corresponding enzymatic characteristics is an important step towards improving the activity, thermostability and even production of heterologous [beta]-glucosidase by glycan engineering. Biotechnol. Bioeng. 2013;110: 3075-3084. [c] 2013 Wiley Periodicals, Inc. Article Note: This article was published online on 1 August 2013. Subsequently, an error was found and the correction was published on 16 August 2013. Wei Wei and Ling Chen contributed equally to this work. Supporting information: Additional Supporting Information may be found in the online version of this article Additional supporting information may be found in the online version of this article at the publisher's web-site. CAPTION(S): Figure S1.The MALDI-TOF mass spectrometry of TrBglS (A), TrN224Q (B), TrN295Q (C), TrN363Q (D) and TrN429Q (E). Figure S2. SDS-PAGE of the fermentation supernatant of T. reesei RC30-8 (Lane 3) and the T. reesei strains expressing TrBglS (Lane 2) and TrN224Q (Lane 1). An equal volume of the culture supernatant was loaded. The band representing BglS is indicated with an arrow. The unrelated lanes were removed for clarity. Table SI. Oligonucleotides used in this study.
language: English
source:
identifier: ISSN: 0006-3592
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  • 0006-3592
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titleN-glycosylation affects the proper folding, enzymatic characteristics and production of a fungal [beta]-glucosidase
creatorWei, Wei ; Chen, Ling ; Zou, Gen ; Wang, Qianfu ; Yan, Xing ; Zhang, Jun ; Wang, Chengshu ; Zhou, Zhihua
ispartofBiotechnology and Bioengineering, Dec, 2013, Vol.110(12), p.3075(10)
identifierISSN: 0006-3592
subjectMonosaccharides ; Enzymes ; Asparagine
descriptionByline: Wei Wei, Ling Chen, Gen Zou, Qianfu Wang, Xing Yan, Jun Zhang, Chengshu Wang, Zhihua Zhou ABSTRACT Heterologous expression of [beta]-glucosidase is one of the approaches to enhance the efficiency of fungal cellulase preparations. It has been reported that N-glycosylation affects the structure framework, function and stability of proteins. In this study, a [beta]-glucosidase from Aspergillus terreus (GenBank: XP_001216552, BglS) was heterologously expressed in Pichia pastoris and Trichoderma reesei. The four asparagine residues were all linked with high-mannose-type oligosaccharides in P. pastoris, whereas only N224 carried high-mannose-type glycan in T. reesei (the other three sites carried one N-acetylglucosamine). The long N-glycan chains on PpBglS weakened its substrate affinity, activity and thermostability. The moderate post-translational and post-secretory glycan modification in T. reesei makes it a suitable expression system for BglS. The N224 glycan played a critical role in BglS folding. The elucidation of the correlation between the different N-glycosylation patterns of BglS and their corresponding enzymatic characteristics is an important step towards improving the activity, thermostability and even production of heterologous [beta]-glucosidase by glycan engineering. Biotechnol. Bioeng. 2013;110: 3075-3084. [c] 2013 Wiley Periodicals, Inc. Article Note: This article was published online on 1 August 2013. Subsequently, an error was found and the correction was published on 16 August 2013. Wei Wei and Ling Chen contributed equally to this work. Supporting information: Additional Supporting Information may be found in the online version of this article Additional supporting information may be found in the online version of this article at the publisher's web-site. CAPTION(S): Figure S1.The MALDI-TOF mass spectrometry of TrBglS (A), TrN224Q (B), TrN295Q (C), TrN363Q (D) and TrN429Q (E). Figure S2. SDS-PAGE of the fermentation supernatant of T. reesei RC30-8 (Lane 3) and the T. reesei strains expressing TrBglS (Lane 2) and TrN224Q (Lane 1). An equal volume of the culture supernatant was loaded. The band representing BglS is indicated with an arrow. The unrelated lanes were removed for clarity. Table SI. Oligonucleotides used in this study.
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titleN-glycosylation affects the proper folding, enzymatic characteristics and production of a fungal [beta]-glucosidase.
descriptionByline: Wei Wei, Ling Chen, Gen Zou, Qianfu Wang, Xing Yan, Jun Zhang, Chengshu Wang, Zhihua Zhou ABSTRACT Heterologous expression of [beta]-glucosidase is one of the approaches to enhance the efficiency of fungal cellulase preparations. It has been reported that N-glycosylation affects the structure framework, function and stability of proteins. In this study, a [beta]-glucosidase from Aspergillus terreus (GenBank: XP_001216552, BglS) was heterologously expressed in Pichia pastoris and Trichoderma reesei. The four asparagine residues were all linked with high-mannose-type oligosaccharides in P. pastoris, whereas only N224 carried high-mannose-type glycan in T. reesei (the other three sites carried one N-acetylglucosamine). The long N-glycan chains on PpBglS weakened its substrate affinity, activity and thermostability. The moderate post-translational and post-secretory glycan modification in T. reesei makes it a suitable expression system for BglS. The N224 glycan played a critical role in BglS folding. The elucidation of the correlation between the different N-glycosylation patterns of BglS and their corresponding enzymatic characteristics is an important step towards improving the activity, thermostability and even production of heterologous [beta]-glucosidase by glycan engineering. Biotechnol. Bioeng. 2013;110: 3075-3084. [c] 2013 Wiley Periodicals, Inc. Article Note: This article was published online on 1 August 2013. Subsequently, an error was found and the correction was published on 16 August 2013. Wei Wei and Ling Chen contributed equally to this work. Supporting information: Additional Supporting Information may be found in the online version of this article Additional supporting information may be found in the online version of this article at the publisher's web-site. CAPTION(S): Figure S1.The MALDI-TOF mass spectrometry of TrBglS (A), TrN224Q (B), TrN295Q (C), TrN363Q (D) and TrN429Q (E). Figure S2. SDS-PAGE of the fermentation supernatant of T. reesei RC30-8 (Lane 3) and the T. reesei strains expressing TrBglS (Lane 2) and TrN224Q (Lane 1). An equal volume of the culture supernatant was loaded. The band representing BglS is indicated with an arrow. The unrelated lanes were removed for clarity. Table SI. Oligonucleotides used in this study.
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abstractByline: Wei Wei, Ling Chen, Gen Zou, Qianfu Wang, Xing Yan, Jun Zhang, Chengshu Wang, Zhihua Zhou ABSTRACT Heterologous expression of [beta]-glucosidase is one of the approaches to enhance the efficiency of fungal cellulase preparations. It has been reported that N-glycosylation affects the structure framework, function and stability of proteins. In this study, a [beta]-glucosidase from Aspergillus terreus (GenBank: XP_001216552, BglS) was heterologously expressed in Pichia pastoris and Trichoderma reesei. The four asparagine residues were all linked with high-mannose-type oligosaccharides in P. pastoris, whereas only N224 carried high-mannose-type glycan in T. reesei (the other three sites carried one N-acetylglucosamine). The long N-glycan chains on PpBglS weakened its substrate affinity, activity and thermostability. The moderate post-translational and post-secretory glycan modification in T. reesei makes it a suitable expression system for BglS. The N224 glycan played a critical role in BglS folding. The elucidation of the correlation between the different N-glycosylation patterns of BglS and their corresponding enzymatic characteristics is an important step towards improving the activity, thermostability and even production of heterologous [beta]-glucosidase by glycan engineering. Biotechnol. Bioeng. 2013;110: 3075-3084. [c] 2013 Wiley Periodicals, Inc. Article Note: This article was published online on 1 August 2013. Subsequently, an error was found and the correction was published on 16 August 2013. Wei Wei and Ling Chen contributed equally to this work. Supporting information: Additional Supporting Information may be found in the online version of this article Additional supporting information may be found in the online version of this article at the publisher's web-site. CAPTION(S): Figure S1.The MALDI-TOF mass spectrometry of TrBglS (A), TrN224Q (B), TrN295Q (C), TrN363Q (D) and TrN429Q (E). Figure S2. SDS-PAGE of the fermentation supernatant of T. reesei RC30-8 (Lane 3) and the T. reesei strains expressing TrBglS (Lane 2) and TrN224Q (Lane 1). An equal volume of the culture supernatant was loaded. The band representing BglS is indicated with an arrow. The unrelated lanes were removed for clarity. Table SI. Oligonucleotides used in this study.
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