Abstract

UNLABELLED

5-Oxo-L-proline (5-OP) is generated by spontaneous cyclization of L-glutamate and L-glutamine and is, therefore, expected to occur in amino acid-rich environments as well as in all cells. However, its utilization as a substrate by bacteria has rarely been examined. Here, we show that the actinobacterium Corynebacterium glutamicum grows efficiently in minimal medium with 5-OP as the sole carbon and nitrogen source. Under these conditions, the pxpTABC gene cluster was essential. pxpT encodes a secondary transporter of the APC superfamily that likely mediates 5-OP uptake, whereas pxpABC encode a recently identified ATP-dependent 5-oxoprolinase that converts 5-OP to L-glutamate for further metabolism. Both PxpT and PxpABC were required for growth on 5-OP. Upstream of and divergently transcribed from pxpT is pxpR, which encodes a GntR-type transcriptional regulator. Deletion of pxpR enhanced growth on 5-OP, suggesting that PxpR represses the expression of the pxpTABC operon. This conclusion was supported by reporter gene analyses and identification of the PxpR-binding site. In addition, isothermal titration calorimetry showed that purified PxpR binds 5-OP with a K D of 726 ± 23 nM. In contrast, L-proline, L-glutamate, L-glutamine, and L-aspartate were not bound under the conditions tested, indicating that PxpR functions as a specific 5-OP sensor. These findings identify a dedicated uptake, catabolic, and regulatory system for 5-OP utilization in C. glutamicum.

IMPORTANCE

Bacterial utilization of 5-oxoproline (5-OP) as a carbon and nitrogen source has received little attention. Because 5-OP forms spontaneously from L-glutamate and L-glutamine, it is likely ubiquitous in amino acid-rich environments as well as inside cells. In many bacteria, including Corynebacterium glutamicum, Escherichia coli, and Bacillus subtilis, intracellular L-glutamate concentrations can reach 100 mM or higher, favoring 5-OP formation. Here, we show that C. glutamicum uses 5-oxoprolinase (PxpABC) together with the transporter PxpT to grow on 5-OP as the sole carbon and nitrogen source. The widespread distribution of pxpABC genes further suggests that the capacity to exploit 5-OP may be common among bacteria and could contribute to growth and survival in diverse habitats.

MeSH

Cells, Proline, Genes, Corynebacterium, Growth, Corynebacterium Glutamicum, Amino Acids, Pyroglutamate Hydrolase, Environment, Bacteria, Glutamine