Abstract
The increasing commercial demand for L-carnitine has led to a multiplication of efforts to improve its production with bacteria. The use of different cell environments, such as growing, resting, permeabilized, dried, osmotically stressed, freely suspended and immobilized cells, to maintain enzymes sufficiently active for L-carnitine production is discussed in the text. The different cell states of enterobacteria, such as Escherichia coli and Proteus sp., which can be used to produce L-carnitine from crotonobetaine or D-carnitine as substrate, are analyzed. Moreover, the combined application of both bioprocess and metabolic engineering has allowed a deeper understanding of the main factors controlling the production process, such as energy depletion and the alteration of the acetyl-CoA/CoA ratio which are coupled to the end of the biotransformation. Furthermore, the profiles of key central metabolic activities such as the TCA cycle, the glyoxylate shunt and the acetate metabolism are seen to be closely interrelated and affect the biotransformation efficiency. Although genetically modified strains have been obtained, new strain improvement strategies are still needed, especially in Escherichia coli as a model organism for molecular biology studies. This review aims to summarize and update the state of the art in L-carnitine production using E. coli and Proteus sp, emphasizing the importance of proper reactor design and operation strategies, together with metabolic engineering aspects and the need for feed-back between wet and in silico work to optimize this biotransformation.
Highlights
In a biotransformation involving whole cells, proper process operation and optimum production depends on the optimization of different variables
The main aim of this review is to update the knowledge on bacterial carnitine metabolism and the potential industrial application of its production methods, focusing on both bioprocess development and strain optimisation strategies
While bioprocesses developed for commercial production of L-carnitine by Sigma Tau (Italy) are based on the biotransformation of crotonobetaine by E. coli and Proteus mirabilis strains, Lonza (Switzerland) uses γ-butyrobetaine as starting material and a derivative of the HK4 strain
Summary
In a biotransformation involving whole cells (either using growing, resting or even permeabilized cells), proper process operation and optimum production depends on the optimization of different variables. We may include (i) the physicochemical conditions of operation for maximum active enzyme concentration per unit of cell (protein synthesis, activity and/ or stability), such as pH, temperature, osmolarity and the presence of activators/inhibitors, (ii) the concentration of both the catalyst and the substrate being used, (iii) the biocatalytic environment (i.e. reactor type), (iv) the optimization scheme, which, in turn, depends on the bioprocess studied, and (v) strain improvement strategies. Enzymes involved in the bioprocess can be either constitutive or induced in certain conditions of cell growth. Process optimization requires a step by step scheme of variable optimization, as will be presented below.
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