Polyelectrolyte Complex Beads by Novel Two-Step Process for Improved Performance of Viable Whole-Cell Baeyer-Villiger Monoxygenase by Immobilization

Tomáš Krajčovič,Vilém Neděla,Igor Lacík,Martin Gericke,Eva Tihlaříková,Marek Bučko,Lucia Uhelská,Dušan Chorvát,Thomas Heinze,Peter Gemeiner,Alica Vikartovská

doi:10.3390/catal7110353

Abstract

A novel immobilization matrix for the entrapment of viable whole-cell Baeyer–Villiger monooxygenase was developed. Viable recombinant Escherichia coli cells overexpressing cyclohexanone monooxygenase were entrapped in polyelectrolyte complex beads prepared by a two-step reaction of oppositely-charged polymers including highly defined cellulose sulphate. Immobilized cells exhibited higher operational stability than free cells during 10 repeated cycles of Baeyer–Villiger biooxidations of rac-bicyclo[3.2.0]hept-2-en-6-one to the corresponding lactones (1R,5S)-3-oxabicyclo-[3.3.0]oct-6-en-3-one and (1S,5R)-2-oxabicyclo-[3.3.0]oct-6-en-3-one. The morphology of polyelectrolyte complex beads was characterised by environmental scanning electron microscopy; the spatial distribution of polymers in the beads and cell viability were examined using confocal laser scanning microscopy, and the texture was characterised by the mechanical resistance measurements.

Highlights

The immobilization of viable whole-cell biocatalysts has long been considered an important method for ensuring their recyclability and stabilisation for the development of the industrial production of chemical specialities and chiral drug precursors [1]
This work focused on the development of an immobilization procedure providing polyelectrolyte complex (PEC) beads for the entrapment of viable recombinant E. coli cells with overexpressed cyclohexanone monooxygenase
Uniform and stable PEC beads were successfully produced by a two-step procedure including ionotropic gelation and polyelectrolyte complexation

Summary

Introduction

The immobilization of viable whole-cell biocatalysts has long been considered an important method for ensuring their recyclability and stabilisation for the development of the industrial production of chemical specialities and chiral drug precursors [1]. One of the validated techniques for immobilizing viable recombinant cells is their encapsulation in polyelectrolyte complex (PEC) capsules, which afforded promising results in the previous development. PA microdrops have been air-stripped by a coaxial nozzle into a polycation solution flowing in a multiloop chemical reactor for the preparation of spherical and monodispersed PEC capsules [7]. This encapsulation protocol preserved the high viability of the encapsulated recombinant Escherichia coli (E. coli) cells with overproduced ezymes from the group of Baeyer–Villiger monooxygenases (BVMOs) as well as their long-term storage and operational stability [4,8,9,10]

Methods

Results

Conclusion