Lysis of a Lactococcus lactis dipeptidase mutant and rescue by mutation in the pleiotropic regulator cody

Chenxi Huang, Jhonatan A Hernandez-Valdes, Oscar P. Kuipers, Jan Kok*

*Corresponding author for this work

Research output: Contribution to journalArticleAcademicpeer-review

2 Citations (Scopus)
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Abstract

Lactococcus lactis subsp. cremoris 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 pepV (creating strain MGΔpepV) resulted in a prolonged lag phase when the mutant strain was grown with a high concentration of glycine. In addition, most MGΔpepV 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 MGΔpepV is more resistant to vancomycin, an antibiotic targeting peptidoglycan D-Ala-D-Ala ends, which confirmed that MGΔpepV has an abnormal PG composition. A mutant of MGΔpepV 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 codY 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 MGΔpepVcodYR218C. Transcriptome sequencing (RNA-seq) data revealed a dramatic derepression in peptide uptake and amino acid utilization in MGΔpepVcodYR218C. A model of the connections among PepV activity, CodY regulation, and PG synthesis of L. lactis is proposed. IMPORTANCE Precise control of peptidoglycan synthesis is essential in Grampositive bacteria for maintaining cell shape and integrity as well as resisting stresses. Although neither the dipeptidase PepV nor alanine is essential for L. lactis 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 L. lactis.

Original languageEnglish
Article number2937
Number of pages15
JournalApplied and environmental microbiology
Volume86
Issue number8
Early online date31-Jan-2020
DOIs
Publication statusPublished - 1-Apr-2020

Keywords

  • Cell morphology
  • Dipeptidase pepV
  • Global repressor CodY
  • Intracellular alanine
  • Lactococcus lactis
  • Nitrogen metabolism
  • Peptidoglycan synthesis

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