Characterization of AtlL, a bifunctional autolysin of Staphylococcus lugdunensis with N-acetylglucosaminidase and N-acetylmuramoyl-l-alanine amidase activities.

Ingrid Bourgeois, Emilie Camiade, Raja Biswas, Pascal Courtin, Laure Gibert, Friedrich Götz, Marie-Pierre Chapot-Chartier, Jean-Louis Pons, Martine Pestel-Caron

Groupe de Recherche sur les Antimicrobiens et les Microorganismes, Rouen University Hospital, Rouen, France.

FEMS microbiology letters 2009 Jan

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The nucleotide sequence of atlL, a gene encoding a putative Staphylococcus lugdunensis peptidoglycan hydrolase, was determined using degenerate consensus PCR and genome walking. This 3837-bp gene encodes a protein, AtlL, that appears as a putative bifunctional autolysin with a 29-amino acid putative signal peptide and two enzymatic putative centres (N-acetylmuramoyl-l-alanine amid ase and N-acetylglucosaminidase) interconnected with three imperfect repeated sequences displaying glycine-tryptophan motifs. In order to determine whether both lytic domains were functional, and verify their exact enzymatic activities, gene fragments harbouring both putative domains, AM (N-acetylmuramoyl-l-alanine amid ase enzymatic centre plus two repeated sequences) and GL (N-acetylglucosaminidase enzymatic centre plus one repeated sequence), were isolated, subcloned, and expressed in Escherichia coli. Purified recombinant AM and GL protein truncations exhibited cell wall lytic activity in zymograms performed with cell walls of Micrococcus lysodeikticus, Bacillus subtilis, and S. lugdunensis. AtlL is expressed during the whole growth, with an overexpression in the early-exponential stage. Liquid chromatography-mass spectrometry analysis of muropeptides generated by digestion of B. subtilis cell walls demonstrated the hydrolytic bond specificities and confirmed both of the acetyl domains' activities as predicted by sequence homology data. AtlL is the first autolysin described in S. lugdunensis, with a bifunctional enzymatic activity involved in peptidoglycan hydrolysis.

Citation

Ingrid Bourgeois, Emilie Camiade, Raja Biswas, Pascal Courtin, Laure Gibert, Friedrich Götz, Marie-Pierre Chapot-Chartier, Jean-Louis Pons, Martine Pestel-Caron. Characterization of AtlL, a bifunctional autolysin of Staphylococcus lugdunensis with N-acetylglucosaminidase and N-acetylmuramoyl-l-alanine amidase activities. FEMS microbiology letters. 2009 Jan;290(1):105-13