Lysis of a Lactococcus lactis Dipeptidase Mutant and Rescue by Mutation in the Pleiotropic Regulator CodY.

subsp. MG1363 is a model for the lactic acid bacteria (LAB) used in the dairy industry. The proteolytic system, consisting of a proteinase, several peptide and amino acid uptake systems, and a host of intracellular peptidases, plays a vital role in nitrogen metabolism and is of eminent importance for flavor formation in dairy products. The dipeptidase PepV functions in the last stages of proteolysis. A link between nitrogen metabolism and peptidoglycan (PG) biosynthesis was underlined by the finding that deletion of the dipeptidase gene (creating strain MGΔ) resulted in a prolonged lag phase when the mutant strain was grown with a high concentration of glycine. In addition, most MGΔ cells lyse and have serious defects in their shape. This phenotype is due to a shortage of alanine, since adding alanine can rescue the growth and shape defects. Strain MGΔ is more resistant to vancomycin, an antibiotic targeting peptidoglycan d-Ala-d-Ala ends, which confirmed that MGΔ has an abnormal PG composition. A mutant of MGΔ was obtained in which growth inhibition and cell shape defects were alleviated. Genome sequencing showed that this mutant has a single point mutation in the gene, resulting in an arginine residue at position 218 in the DNA-binding motif of CodY being replaced by a cysteine residue. Thus, this strain was named MGΔ Transcriptome sequencing (RNA-seq) data revealed a dramatic derepression in peptide uptake and amino acid utilization in MGΔ A model of the connections among PepV activity, CodY regulation, and PG synthesis of is proposed. Precise control of peptidoglycan synthesis is essential in Gram-positive bacteria for maintaining cell shape and integrity as well as resisting stresses. Although neither the dipeptidase PepV nor alanine is essential for MG1363, adequate availability of either ensures proper cell wall synthesis. We broaden the knowledge about the dipeptidase PepV, which acts as a linker between nitrogen metabolism and cell wall synthesis in .