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A simplified process design for P450 driven hydroxylation based on surface displayed enzymes.

Frank W Ströhle1, Eva Kranen2, Jens Schrader1

  • 1DECHEMA-Forschungsinstitut, Biochemical Engineering, Frankfurt am Main, Hessen, 60486, Germany.

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Summary
This summary is machine-generated.

Researchers engineered E. coli to display P450BM3 enzymes on their surface for sustainable chemical production. This biocatalyst efficiently hydroxylates fatty acids and can be reused without activity loss, achieving high turnover numbers.

Keywords:
P450 monooxygenasescofactor regenerationprocess designsurface displaytotal turnover numbers

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Area of Science:

  • Biocatalysis and metabolic engineering
  • Sustainable chemical synthesis
  • Protein engineering

Background:

  • Developing sustainable industrial chemical production requires optimizing existing processes and improving catalyst utilization.
  • Surface display of enzymes offers a promising strategy for enhanced biocatalyst performance and reusability.

Purpose of the Study:

  • To express P450BM3 on the surface of E. coli using the Autodisplay system.
  • To evaluate cofactor regeneration systems and develop a reusable biocatalyst for fatty acid hydroxylation.
  • To achieve high turnover numbers for P450 monooxygenases in industrial applications.

Main Methods:

  • Engineered E. coli for surface expression of P450BM3 via the Autodisplay system.
  • Investigated and compared various enzymatic cofactor regeneration systems.
  • Developed a process for repeated use of surface-displayed biocatalysts after simple centrifugation.

Main Results:

  • Successfully demonstrated surface expression of P450BM3 and its activity in palmitic acid hydroxylation.
  • The biocatalyst showed no loss of activity after multiple reuses.
  • Achieved a total turnover number of up to 54,700, a record for P450 monooxygenases, using surface-displayed P450s with enzymatic cofactor regeneration.

Conclusions:

  • Surface display of P450BM3 on E. coli is an effective strategy for sustainable biocatalysis.
  • The developed system enables efficient and reusable biocatalytic hydroxylation of fatty acids.
  • This approach holds significant potential for advancing sustainable bioprocess design in the chemical industry.