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Applied Microbiology and Biotechnology

, Volume 102, Issue 12, pp 5185–5196 | Cite as

Evolution of the feruloyl esterase MtFae1a from Myceliophthora thermophila towards improved catalysts for antioxidants synthesis

  • Simona Varriale
  • Gabriella Cerullo
  • Io Antonopoulou
  • Paul Christakopoulos
  • Ulrika Rova
  • Thierry Tron
  • Régis Fauré
  • Peter Jütten
  • Alexander Piechot
  • Joana L. A. Brás
  • Carlos M. G. A. Fontes
  • Vincenza Faraco
Biotechnologically relevant enzymes and proteins

Abstract

The chemical syntheses currently employed for industrial purposes, including in the manufacture of cosmetics, present limitations such as unwanted side reactions and the need for harsh chemical reaction conditions. In order to overcome these drawbacks, novel enzymes are developed to catalyze the targeted bioconversions. In the present study, a methodology for the construction and the automated screening of evolved variants library of a Type B feruloyl esterase from Myceliophthora thermophila (MtFae1a) was developed and applied to generation of 30,000 mutants and their screening for selecting the variants with higher activity than the wild-type enzyme. The library was generated by error-prone PCR of mtfae1a cDNA and expressed in Saccharomyces cerevisiae. Screening for extracellular enzymatic activity towards 4-nitrocatechol-1-yl ferulate, a new substrate developed ad hoc for high-throughput assays of feruloyl esterases, led to the selection of 30 improved enzyme variants. The best four variants and the wild-type MtFae1a were investigated in docking experiments with hydroxycinnamic acid esters using a model of 3D structure of MtFae1a. These variants were also used as biocatalysts in transesterification reactions leading to different target products in detergentless microemulsions and showed enhanced synthetic activities, although the screening strategy had been based on improved hydrolytic activity.

Keywords

Directed evolution High-throughput screening Myceliophthora thermophila Library Feruloyl esterase 

Notes

Acknowledgements

This work was supported by a grant from the European Union—Large-scale integrating project targeted to SMEs “Optimized esterase biocatalysts for cost-effective industrial production (OPTIBIOCAT)” grant agreement no. 613868, co-funded within the FP7 Knowledge Based Bio-Economy (KBBE).

Funding

This study was funded by grant from the European Union Grant agreement no. 613868.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

Ethical approval

This article does not contain any studies with human participants or animals performed by any of the authors.

Supplementary material

253_2018_8995_MOESM1_ESM.pdf (145 kb)
ESM 1 (PDF 144 kb)

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Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2018

Authors and Affiliations

  • Simona Varriale
    • 1
  • Gabriella Cerullo
    • 1
  • Io Antonopoulou
    • 2
  • Paul Christakopoulos
    • 2
  • Ulrika Rova
    • 2
  • Thierry Tron
    • 3
  • Régis Fauré
    • 4
  • Peter Jütten
    • 5
  • Alexander Piechot
    • 5
  • Joana L. A. Brás
    • 6
  • Carlos M. G. A. Fontes
    • 6
  • Vincenza Faraco
    • 1
  1. 1.Department of Chemical Sciences, Complesso Universitario Monte S. AngeloUniversity of Naples Federico IINaplesItaly
  2. 2.Biochemical Process Engineering, Division of Chemical Engineering, Department of Civil, Environmental and Natural Resources EngineeringLuleå University of TechnologyLuleåSweden
  3. 3.Centrale Marseille, CNRSAix Marseille UniversitéMarseilleFrance
  4. 4.LISBP, CNRS, INRA, INSAUniversité de ToulouseToulouseFrance
  5. 5.Taros Chemicals GmbH & Co. KGDortmundGermany
  6. 6.NzyTech LDALisbonPortugal

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