Ethambutol-mediated cell wall modification in recombinant Corynebacterium glutamicum increases the biotransformation rates of cyclohexanone derivatives

The effects of structural modification of cell wall on the biotransformation capability by recombinant Corynebacterium glutamicum cells, expressing the chnB gene encoding cyclohexanone monooxygenase of Acinetobacter calcoaceticus NCIMB 9871, were investigated. Baeyer-Villiger oxygenation of 2-(20-acetoxyethyl) cyclohexanone (MW 170 Da) into R-7-(20-acetoxyethyl)-2- oxepanone was used as a model reaction. The whole-cell biotransformation followed Michaelis-Menten kinetics. The Vmax and KS values were estimated as 96.8 U g-1 of dry cells and 0.98 mM, respectively. The Vmax was comparable with that of cyclohexanone oxygenation, whereas the KS was almost eightfold higher. The KS value of 2-(20-acetoxyethyl) cyclohexanone oxygenation was reduced by ca. 30% via altering the cell envelop structure of C. glutamicum with ethambutol, which inhibits arabinosyl transferases involved in the biosynthesis of cell wall arabinogalactan and mycolate layers. The higher wholecell biotransformation rate was also observed in the oxygenation of ethyl 2-cyclohexanone acetate upon ethambutol treatment of the recombinant C. glutamicum. Therefore, it was assumed that the biotransformation efficiency of C. glutamicum-based biocatalysts, with respect to medium- to large-sized lipophilic organic substrates (MW[ca. 170), can be enhanced by engineering their cell wall outer layers, which are known to function as a formidable barrier to lipophilic molecules.