Immobilization of Escherichia coli cells expressing 4-oxalocrotonate tautomerase for improved biotransformation of β-nitrostyrene

Abstract

The enzyme 4-oxalocrotonate tautomerase (4-OT) encoded by the xylH gene is a part of the degradation pathway of aromatic compounds in Pseudomonas putida mt-2. 4-OT was described to catalyze Michael-type addition of acetaldehyde to β-nitrostyrene, and the whole cell system based on recombinantly expressed 4-OT has been developed previously. In this study biocatalytic process based on Escherichia coli whole cells expressing 4-OT was significantly improved using immobilization and ex situ product recovery strategies. Whole cell immobilization in alginate beads was applied in biocatalytic production of 4-nitro-3-phenyl-butanal from β-nitrostyrene and acetaldehyde. Immobilized biocatalyst showed wider pH activity range and could tolerate twofold higher initial concentrations of substrate in comparison to the free whole cell biocatalyst. Beads retained their initial activity over 10 consecutive biotransformations of the model reaction and remained suitable for the repetitive use with 85% of the initial activity after two months of storage. Bioprocess was further improved by utilizing Amberlite XAD-2 hydrophobic resin for the product recovery. With this modification, the amount of organic solvent was reduced 40-fold in comparison to previously reported method making this biocatalytic process greener.

Improvement of caffeic acid biotransformation into para‐hydroxybenzoic acid by Candida albicans CI‐24 via gamma irradiation and model‐based optimization

“…Similarly, Velankar et al 39 has reported that alginate-immobilized Rhodotorula minuta cells could biotransform citronellal for three successive runs. Interestingly, Djokic et al 40 immobilized E. coli cells that were able to transform βnitrostyrene for 11 consecutive cycles.…”

Enabling protein-hosted organocatalytic transformations

“…4-OT and its variants have been used for a range of applications including enzymatic 115 and chemoenzymatic cascades, 116 alongside whole cell catalytic systems. [117][118][119] The anti-anxiety drug pregabalin and three of its analogues were synthesised by coupling the 4-OT reaction with catalysis by aldehyde dehydrogenase (ALDH, Fig. 8).…”

Characterization of Functional Biohybrid Materials Based on Saccharomyces Cerevisiae Biomass

“…[5] Origins of materials shaped by microorganisms can be seen in whole-cell biocatalysis: encapsulation of cells into hydrogel matrices such as κ-carrageenan, Ca-alginate, polyvinyl alcohol, and polyacrylamide was used to improve the catalytic efficiency, catalyst lifetime, and reusability. [6][7][8][9] Functional coatings have been obtained by doping latex coatings with low amounts of microorganisms (<20% w/w) which exhibited photosynthetic, oxidative, or hydrolytic activity. [10][11][12] Liu et al reported stimuliresponsive protein expression within 3D-printable hydrogels.…”