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Molecular characterization of the thermostability and carbohydrate-binding module from a newly identified GH118 family agarase, AgaXa.

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Title: Molecular characterization of the thermostability and carbohydrate-binding module from a newly identified GH118 family agarase, AgaXa.
Authors: Wu, Yi-Rui1, Zhou, Zheng-Rong1, Zhao, Min1, Lin, Bokun1, Zhong, Mingqi1, Hu, Zhong1 hzh@stu.edu.cn
Source: Process Biochemistry. Jan2017, Vol. 52, p192-199. 8p.
Subjects: Carbohydrates, Thermal stability, Protein stability, Glycosidases, Circular dichroism, Site-specific mutagenesis, Ligand binding (Biochemistry)
Abstract: AgaXa is a thermostable β-agarase from the agar-degrading bacterium Catenovulum sp. X3. To further understand the mechanism of protein stabilization of AgaXa, several mutants were generated by random and site-directed mutagenesis, and they were subsequently screened by analysing their enzymatic activity and thermostability. Four mutants (V197D, P259H, C442S and C528S) were found for which the enzyme activity and thermostability were significantly decreased. Moreover, secondary structures determined by circular dichroism (CD) showed that mutants V197D and P259H presented a higher percentage of α-helix but fewer β-strands than wild-type (WT) AgaXa. On the contrary, no significant changes were observed in the secondary structures of mutants C442S and C528S. Through the treatment by proteinase K, different digested profiles were generated from mutants V197D and P259H when compared to WT, and mutants C442S and C528S was observed with more digested protein fragments. These results indicate that the enzymatic activity and stability of AgaXa is mainly related to its hydrophobic structure and disulphide bonds. Furthermore, carbohydrate-binding ability was also analysed for the mutants of N- and C-terminal deletions, and the results showed that agarose binding capability was not changed, indicating that the carbohydrate-binding module is probably located in the middle of the β-agarase AgaXa. [ABSTRACT FROM AUTHOR]
Copyright of Process Biochemistry is the property of Elsevier B.V. and its content may not be copied or emailed to multiple sites without the copyright holder's express written permission. Additionally, content may not be used with any artificial intelligence tools or machine learning technologies. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)
Database: Engineering Source
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Header DbId: egs
DbLabel: Engineering Source
An: 120560300
AccessLevel: 6
PubType: Academic Journal
PubTypeId: academicJournal
PreciseRelevancyScore: 0
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  Label: Title
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  Data: Molecular characterization of the thermostability and carbohydrate-binding module from a newly identified GH118 family agarase, AgaXa.
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  Data: <searchLink fieldCode="AR" term="%22Wu%2C+Yi-Rui%22">Wu, Yi-Rui</searchLink><relatesTo>1</relatesTo><br /><searchLink fieldCode="AR" term="%22Zhou%2C+Zheng-Rong%22">Zhou, Zheng-Rong</searchLink><relatesTo>1</relatesTo><br /><searchLink fieldCode="AR" term="%22Zhao%2C+Min%22">Zhao, Min</searchLink><relatesTo>1</relatesTo><br /><searchLink fieldCode="AR" term="%22Lin%2C+Bokun%22">Lin, Bokun</searchLink><relatesTo>1</relatesTo><br /><searchLink fieldCode="AR" term="%22Zhong%2C+Mingqi%22">Zhong, Mingqi</searchLink><relatesTo>1</relatesTo><br /><searchLink fieldCode="AR" term="%22Hu%2C+Zhong%22">Hu, Zhong</searchLink><relatesTo>1</relatesTo><i> hzh@stu.edu.cn</i>
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  Data: <searchLink fieldCode="JN" term="%22Process+Biochemistry%22">Process Biochemistry</searchLink>. Jan2017, Vol. 52, p192-199. 8p.
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  Data: <searchLink fieldCode="DE" term="%22Carbohydrates%22">Carbohydrates</searchLink><br /><searchLink fieldCode="DE" term="%22Thermal+stability%22">Thermal stability</searchLink><br /><searchLink fieldCode="DE" term="%22Protein+stability%22">Protein stability</searchLink><br /><searchLink fieldCode="DE" term="%22Glycosidases%22">Glycosidases</searchLink><br /><searchLink fieldCode="DE" term="%22Circular+dichroism%22">Circular dichroism</searchLink><br /><searchLink fieldCode="DE" term="%22Site-specific+mutagenesis%22">Site-specific mutagenesis</searchLink><br /><searchLink fieldCode="DE" term="%22Ligand+binding+%28Biochemistry%29%22">Ligand binding (Biochemistry)</searchLink>
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  Label: Abstract
  Group: Ab
  Data: AgaXa is a thermostable β-agarase from the agar-degrading bacterium Catenovulum sp. X3. To further understand the mechanism of protein stabilization of AgaXa, several mutants were generated by random and site-directed mutagenesis, and they were subsequently screened by analysing their enzymatic activity and thermostability. Four mutants (V197D, P259H, C442S and C528S) were found for which the enzyme activity and thermostability were significantly decreased. Moreover, secondary structures determined by circular dichroism (CD) showed that mutants V197D and P259H presented a higher percentage of α-helix but fewer β-strands than wild-type (WT) AgaXa. On the contrary, no significant changes were observed in the secondary structures of mutants C442S and C528S. Through the treatment by proteinase K, different digested profiles were generated from mutants V197D and P259H when compared to WT, and mutants C442S and C528S was observed with more digested protein fragments. These results indicate that the enzymatic activity and stability of AgaXa is mainly related to its hydrophobic structure and disulphide bonds. Furthermore, carbohydrate-binding ability was also analysed for the mutants of N- and C-terminal deletions, and the results showed that agarose binding capability was not changed, indicating that the carbohydrate-binding module is probably located in the middle of the β-agarase AgaXa. [ABSTRACT FROM AUTHOR]
– Name: AbstractSuppliedCopyright
  Label:
  Group: Ab
  Data: <i>Copyright of Process Biochemistry is the property of Elsevier B.V. and its content may not be copied or emailed to multiple sites without the copyright holder's express written permission. Additionally, content may not be used with any artificial intelligence tools or machine learning technologies. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract.</i> (Copyright applies to all Abstracts.)
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RecordInfo BibRecord:
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      – Type: doi
        Value: 10.1016/j.procbio.2016.10.021
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      – Code: eng
        Text: English
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        PageCount: 8
        StartPage: 192
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      – SubjectFull: Carbohydrates
        Type: general
      – SubjectFull: Thermal stability
        Type: general
      – SubjectFull: Protein stability
        Type: general
      – SubjectFull: Glycosidases
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      – SubjectFull: Circular dichroism
        Type: general
      – SubjectFull: Site-specific mutagenesis
        Type: general
      – SubjectFull: Ligand binding (Biochemistry)
        Type: general
    Titles:
      – TitleFull: Molecular characterization of the thermostability and carbohydrate-binding module from a newly identified GH118 family agarase, AgaXa.
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            NameFull: Wu, Yi-Rui
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            NameFull: Zhou, Zheng-Rong
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            NameFull: Zhao, Min
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            NameFull: Lin, Bokun
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            NameFull: Zhong, Mingqi
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              M: 01
              Text: Jan2017
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              Y: 2017
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              Value: 52
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            – TitleFull: Process Biochemistry
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