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Cell Surface Display of Poly(3-hydroxybutyrate) Depolymerase and its Application.

Seung Hwan Lee1, Sang Yup Lee2

  • 1Department of Biotechnology and Bioengineering, Chonnam National University, Gwangju 61186, Republic of Korea.

Journal of Microbiology and Biotechnology
|February 19, 2020
PubMed
Summary
This summary is machine-generated.

We engineered Escherichia coli to display poly(3-hydroxybutyrate) (PHB) depolymerase on its surface. This surface-displayed enzyme acts as an efficient biocatalyst for enantioselective hydrolysis, producing high-purity mandelic acid.

Keywords:
Cell surface displayOprFdepolymeraseenantioselective biocatalysimmobilizationwhole cell biocatalyst

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Area of Science:

  • Biotechnology
  • Enzyme Engineering
  • Microbial Surface Display

Background:

  • Extracellular poly(3-hydroxybutyrate) (PHB) depolymerase from Ralstonia pickettii T1 is a valuable enzyme.
  • Efficient methods for enzyme immobilization and application are needed for biocatalysis.
  • Surface display of enzymes on microbial hosts offers advantages for catalytic applications.

Purpose of the Study:

  • To express and display Ralstonia pickettii T1 PHB depolymerase on the surface of Escherichia coli.
  • To evaluate the potential of surface-displayed depolymerase as an immobilized biocatalyst for enantioselective hydrolysis.
  • To demonstrate the application of this system in the synthesis of chiral compounds.

Main Methods:

  • C-terminal deletion-fusion strategy using Pseudomonas OprF protein as a fusion partner.
  • Surface display confirmation via flow cytometry, immunofluorescence microscopy, and whole-cell hydrolase activity assays.
  • Application as an immobilized catalyst for the enantioselective hydrolysis of methyl mandelate.

Main Results:

  • Successful surface display of active PHB depolymerase on Escherichia coli was confirmed.
  • The immobilized enzyme efficiently catalyzed the enantioselective hydrolysis of (R)-methyl mandelate.
  • (S)-mandelic acid was produced with over 99% enantiomeric excess after 48 hours.

Conclusions:

  • Surface-displayed PHB depolymerase on Escherichia coli is a viable and effective biocatalyst.
  • This system demonstrates potential for industrial applications in enantioselective synthesis.
  • Microbial surface display provides a novel platform for enzyme immobilization and biocatalysis.