Corynebacterium glutamicum as a host for synthesis and export of D-amino acids

作者：Stabler N; Oikawa T; Bott M; Eggeling L^*

来源：Journal of Bacteriology, 2011, 193(7): 1702-1709.

DOI：10.1128/JB.01295-10

A number of D-amino acids occur in nature, and there is growing interest in their function and metabolism, as well as in their production and use. Here we use the well-established L-amino-acid-producing bacterium Corynebacterium glutamicum to study whether D-amino acid synthesis is possible and whether mechanisms for the export of these amino acids exist. In contrast to Escherichia coli, C. glutamicum tolerates D-amino acids added extracellularly. Expression of argR (encoding the broad-substrate-specific racemase of Pseudomonas taetrolens) with its signal sequence deleted results in cytosolic localization of ArgR in C. glutamicum. The isolated enzyme has the highest activity with lysine (100%) but also exhibits activity with serine (2%). Upon overexpression of argR in an L-arginine, L-ornithine, or L-lysine producer, equimolar mixtures of the D- and L-enantiomers accumulated extracellularly. Unexpectedly, argR overexpression in an L-serine producer resulted in extracellular accumulation of a surplus of D-serine (81 mM D-serine and 37 mM L-serine) at intracellular concentrations of 125 mM D-serine plus 125 mM L-serine. This points to a nonlimiting ArgR activity for intracellular serine racemization and to the existence of a specific export carrier for D-serine. Export of D-lysine relies fully on the presence of lysE, encoding the exporter for L-lysine, which is apparently promiscuous with respect to the chirality of lysine. These data show that D-amino acids can also be produced with C. glutamicum and that in special cases, due to specific carriers, even a preferential extracellular accumulation of this enantiomer is possible.

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