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The MurC ligase essential for peptidoglycan biosynthesis is regulated by the serine/threonine protein kinase PknA in $Corynebacterium\ glutamicum$
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Edité par CCSD ; American Society for Biochemistry and Molecular Biology -
International audience. The Mur ligases play an essential role in the biosynthesis ofbacterial cell-wall peptidoglycan and thus represent attractivetargets for the design of novel antibacterials. These enzymescatalyze the stepwise formation of the peptide moiety of thepeptidoglycan disaccharide peptide monomer unit. MurC isresponsible of the addition of the first residue (L-alanine) ontothe nucleotide precursor UDP-MurNAc. Phosphorylation ofproteins by Ser/Thr protein kinases has recently emerged as amajor physiological mechanism of regulation in prokaryotes.Herein, the hypothesis of a phosphorylation-dependent mechanism of regulation of the MurC activity was investigated inCorynebacterium glutamicum. We showed that MurC wasphosphorylated in vitro by the PknA protein kinase. An analysisof the phosphoamino acid content indicated that phosphorylation exclusively occurred on threonine residues. Six phosphoacceptor residues were identified by mass spectrometry analysis,and we confirmed that mutagenesis to alanine residues totallyabolished PknA-dependent phosphorylation of MurC. In vitroand in vivo ligase activity assays showed that the catalytic activity of MurC was impaired following mutation of these threonineresidues. Further in vitro assays revealed that the activity of theMurC-phosphorylated isoform was severely decreased compared with the non-phosphorylated protein. To our knowledge,this is the first demonstration of a MurC ligase phosphorylationin vitro. The finding that phosphorylation is correlated with adecrease in MurC enzymatic activity could have significant consequences in the regulation of peptidoglycan biosynthesis.
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