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DMSO-induced structural and catalytic changes in C-type halohydrin dehalogenase

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Title: DMSO-induced structural and catalytic changes in C-type halohydrin dehalogenase
Authors: Milčić, Nevena, Stepanić, Višnja, Crnolatac, Ivo, Findrik Blažević, Zvjezdana, Brkljača, Zlatko, Majerić-Elenkov, Maja
Publisher Information: 2022.
Publication Year: 2022
Subject Terms: halohydrin dehalogenases, stability, DMSO, molecular dynamics, co-solvent, enzyme activity
Description: Halohydrin dehalogenases (HHDHs) are an important group of enzymes for the synthesis of low molecular weight building blocks with high optical purity. Although they naturally catalyze the reversible dehalogenation of vicinal haloalcohols, their distinctive synthetic and practical feature is ability to utilize a whole range of unnatural nucleophiles in epoxide ring-opening reactions, thus giving access to new chemical bonds (C-C, C-O, C-S, C-N, etc.). One of the mayor problems concerning HHDH-biocatalytic reactions is poor solubility of substrates, i.e., epoxides, and their susceptibility to hydrolytic decomposition. Biocatalysis is limited to the millimolar scale unless a solvent is introduced into the aqueous reaction medium, which could increase the substrate solubility and availability to the enzyme. Most frequently employed solvent while screening new HHDH-activities or performing established reactions is dimethyl sulfoxide (DMSO) on account of its amphipathic, non-toxic, and recyclable nature. Usually, DMSO is used in volume ratios up to 5% ; however, the resulting solubility increase is still too low for industrial applications. Higher solvent quantities are prerequisite for increasing substrate availability and product yield. On the other hand, elevated DMSO ratios could reflect on enzyme structural properties and catalytic performances. In our study, we demonstrated that DMSO affects HheC catalytic properties in a concentration-dependent manner and in relation to structural changes. We assessed DMSO-induced changes at various levels, by means of kinetic and calorimetric measurements, dynamic light scattering, molecular docking and molecular dynamic (MD) simulations. By combining detailed experimental approach with extensive MD calculations, the underlying mechanisms for the loss of HheC activity in the presence of DMSO were explained. Kinetic measurements showed that co-solvent DMSO is a strong mixed catalytic inhibitor with dominant competitive contribution in chosen reaction, which was further confirmed by MD simulations. The computations showed that DMSO has high predisposition for binding to the Ser-Tyr-Arg catalytic triad. Besides inhibitory properties which occur already at lower DMSO concentrations, structural changes which lead to inactivation and aggregation are dominant at amounts above 30% (v/v). This is the evidence of high robustness and resistance of HheC enzyme against polar solvent action. In conclusion, DMSO displays diversified unfavorable effects on HheC, which include mixed enzyme inhibition, enzyme dehydration, structural distortions and aggregation. These results emphasize the importance of careful solvent type and amount selection in HHDH-catalyzed reactions.
Document Type: Conference object
Accession Number: edsair.dris...01492..3d66cd20c7d7df1ee21596232f0aa6aa
Database: OpenAIRE
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Items – Name: Title
  Label: Title
  Group: Ti
  Data: DMSO-induced structural and catalytic changes in C-type halohydrin dehalogenase
– Name: Author
  Label: Authors
  Group: Au
  Data: <searchLink fieldCode="AR" term="%22Milčić%2C+Nevena%22">Milčić, Nevena</searchLink><br /><searchLink fieldCode="AR" term="%22Stepanić%2C+Višnja%22">Stepanić, Višnja</searchLink><br /><searchLink fieldCode="AR" term="%22Crnolatac%2C+Ivo%22">Crnolatac, Ivo</searchLink><br /><searchLink fieldCode="AR" term="%22Findrik+Blažević%2C+Zvjezdana%22">Findrik Blažević, Zvjezdana</searchLink><br /><searchLink fieldCode="AR" term="%22Brkljača%2C+Zlatko%22">Brkljača, Zlatko</searchLink><br /><searchLink fieldCode="AR" term="%22Majerić-Elenkov%2C+Maja%22">Majerić-Elenkov, Maja</searchLink>
– Name: Publisher
  Label: Publisher Information
  Group: PubInfo
  Data: 2022.
– Name: DatePubCY
  Label: Publication Year
  Group: Date
  Data: 2022
– Name: Subject
  Label: Subject Terms
  Group: Su
  Data: <searchLink fieldCode="DE" term="%22halohydrin+dehalogenases%22">halohydrin dehalogenases</searchLink><br /><searchLink fieldCode="DE" term="%22stability%22">stability</searchLink><br /><searchLink fieldCode="DE" term="%22DMSO%22">DMSO</searchLink><br /><searchLink fieldCode="DE" term="%22molecular+dynamics%22">molecular dynamics</searchLink><br /><searchLink fieldCode="DE" term="%22co-solvent%22">co-solvent</searchLink><br /><searchLink fieldCode="DE" term="%22enzyme+activity%22">enzyme activity</searchLink>
– Name: Abstract
  Label: Description
  Group: Ab
  Data: Halohydrin dehalogenases (HHDHs) are an important group of enzymes for the synthesis of low molecular weight building blocks with high optical purity. Although they naturally catalyze the reversible dehalogenation of vicinal haloalcohols, their distinctive synthetic and practical feature is ability to utilize a whole range of unnatural nucleophiles in epoxide ring-opening reactions, thus giving access to new chemical bonds (C-C, C-O, C-S, C-N, etc.). One of the mayor problems concerning HHDH-biocatalytic reactions is poor solubility of substrates, i.e., epoxides, and their susceptibility to hydrolytic decomposition. Biocatalysis is limited to the millimolar scale unless a solvent is introduced into the aqueous reaction medium, which could increase the substrate solubility and availability to the enzyme. Most frequently employed solvent while screening new HHDH-activities or performing established reactions is dimethyl sulfoxide (DMSO) on account of its amphipathic, non-toxic, and recyclable nature. Usually, DMSO is used in volume ratios up to 5% ; however, the resulting solubility increase is still too low for industrial applications. Higher solvent quantities are prerequisite for increasing substrate availability and product yield. On the other hand, elevated DMSO ratios could reflect on enzyme structural properties and catalytic performances. In our study, we demonstrated that DMSO affects HheC catalytic properties in a concentration-dependent manner and in relation to structural changes. We assessed DMSO-induced changes at various levels, by means of kinetic and calorimetric measurements, dynamic light scattering, molecular docking and molecular dynamic (MD) simulations. By combining detailed experimental approach with extensive MD calculations, the underlying mechanisms for the loss of HheC activity in the presence of DMSO were explained. Kinetic measurements showed that co-solvent DMSO is a strong mixed catalytic inhibitor with dominant competitive contribution in chosen reaction, which was further confirmed by MD simulations. The computations showed that DMSO has high predisposition for binding to the Ser-Tyr-Arg catalytic triad. Besides inhibitory properties which occur already at lower DMSO concentrations, structural changes which lead to inactivation and aggregation are dominant at amounts above 30% (v/v). This is the evidence of high robustness and resistance of HheC enzyme against polar solvent action. In conclusion, DMSO displays diversified unfavorable effects on HheC, which include mixed enzyme inhibition, enzyme dehydration, structural distortions and aggregation. These results emphasize the importance of careful solvent type and amount selection in HHDH-catalyzed reactions.
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  Label: Document Type
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  Data: Conference object
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  Data: edsair.dris...01492..3d66cd20c7d7df1ee21596232f0aa6aa
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RecordInfo BibRecord:
  BibEntity:
    Languages:
      – Text: Undetermined
    Subjects:
      – SubjectFull: halohydrin dehalogenases
        Type: general
      – SubjectFull: stability
        Type: general
      – SubjectFull: DMSO
        Type: general
      – SubjectFull: molecular dynamics
        Type: general
      – SubjectFull: co-solvent
        Type: general
      – SubjectFull: enzyme activity
        Type: general
    Titles:
      – TitleFull: DMSO-induced structural and catalytic changes in C-type halohydrin dehalogenase
        Type: main
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      – PersonEntity:
          Name:
            NameFull: Milčić, Nevena
      – PersonEntity:
          Name:
            NameFull: Stepanić, Višnja
      – PersonEntity:
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            NameFull: Crnolatac, Ivo
      – PersonEntity:
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            NameFull: Findrik Blažević, Zvjezdana
      – PersonEntity:
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            NameFull: Brkljača, Zlatko
      – PersonEntity:
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            NameFull: Majerić-Elenkov, Maja
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          Dates:
            – D: 01
              M: 01
              Type: published
              Y: 2022
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