Factors affecting the biotransformation of trimethylammonium compounds into L-carnitine by Escherichia coli

摘要

The biotransformation of D-carnitine and crotonobetaine into L-carnitine with wild and transformed E.coli strains under batch and continuous operation was optimised.In batch,the best conditions for the transformed strain were 30% oxygen saturation,a temperature of 41 deg C and a minimal medium,whereas anaerobic cultures in either complex or minimal media at 37 deg C and pH 7.5 were optimal for the wild strain.Studies on the expression of the enzymes involved in trimethylammonium metabolism showed that L-carnitine dehydratase activity was always higher than that of D-carnitine racemase.Experiments with the transformed strain in continuous cell-recycle reactors showed that,despite the higher productivity that could be achieved(0.65-1.2 g/L h),plasmid-bearing cells were segregated even when a selective medium was used.This fact was also confirmed by studying the evolution of the D-carnitine racemase level.Immobilization of the transformed strain in K-carrageenan gels allowed continuous operation for L-carnitine production with no plasmid loss.In continuous processes with cell-retention systems,the wild strain showed higher productivity and stability than the transformed strain.Moreover,crotonobetaine was a better substrate for both strains used.Recycling with hollow-fiber cartridges provided the highest biomass level even though the L-carnitine dehydratase/biomass ratio was lower.However,membrane composition and cut-off had less influence on reactor performance as similar levels of productivity were attained.In spite of this,continuous processes attained a L-carnitine production as high as 11.5 g/L h as a result of the high enzyme induction and biomass levels.