title: Specificity of L,D-transpeptidases from gram-positive bacteria producing different peptidoglycan chemotypes.
abstract: We report here the first direct assessment of the specificity of a class of peptidoglycan cross-linking enzymes, the L,D-transpeptidases, for the highly diverse structure of peptidoglycan precursors of Gram-positive bacteria. The lone functionally characterized member of this new family of active site cysteine peptidases, Ldt(fm) from Enterococcus faecium, was previously shown to bypass the D,D-transpeptidase activity of the classical penicillin-binding proteins leading to high level cross-resistance to glycopeptide and beta-lactam antibiotics. Ldt(fm) homologues from Bacillus subtilis (Ldt(Bs)) and E. faecalis (Ldt(fs)) were found here to cross-link their cognate disaccharide-peptide subunits containing meso-diaminopimelic acid (mesoDAP(3)) and L-Lys(3)-L-Ala-L-Ala at the third position of the stem peptide, respectively, instead of L-Lys(3)-d-iAsn in E. faecium. Ldt(fs) differed from Ldt(fm) and Ldt(Bs) by its capacity to hydrolyze the L-Lys(3)-D-Ala(4) bond of tetrapeptide (L,D-carboxypeptidase activity) and pentapeptide (L,D-endopeptidase activity) stems, in addition to the common cross-linking activity. The three enzymes were specific for their cognate acyl acceptors in the cross-linking reaction. In contrast to Ldt(fs), which was also specific for its cognate acyl donor, Ldt(fm) tolerated substitution of L-Lys(3)-D-iAsn by L-Lys(3)-L-Ala-L-Ala. Likewise, Ldt(Bs) tolerated substitution of mesoDAP(3) by L-Lys(3)-D-iAsn and L-Lys(3)-L-Ala-L-Ala in the acyl donor. Thus, diversification of the structure of peptidoglycan precursors associated with speciation has led to a parallel evolution of the substrate specificity of the L,D-transpeptidases affecting mainly the recognition of the acyl acceptor. Blocking the assembly of the side chain could therefore be used to combat antibiotic resistance involving L,D-transpeptidases.
authors.affiliation: INSERM, U655-LRMA F-75006 Paris France.
authors.forename: Sophie
authors.initials: S
authors.lastname: Magnet
authors.forename: Ana
authors.initials: A
authors.lastname: Arbeloa
authors.forename: Jean-Luc
authors.initials: JL
authors.lastname: Mainardi
authors.forename: Jean-Emmanuel
authors.initials: JE
authors.lastname: Hugonnet
authors.forename: Martine
authors.initials: M
authors.lastname: Fourgeaud
authors.forename: Lionel
authors.initials: L
authors.lastname: Dubost
authors.forename: Arul
authors.initials: A
authors.lastname: Marie
authors.forename: Vanessa
authors.initials: V
authors.lastname: Delfosse
authors.forename: Claudine
authors.initials: C
authors.lastname: Mayer
authors.forename: Louis B
authors.initials: LB
authors.lastname: Rice
authors.forename: Michel
authors.initials: M
authors.lastname: Arthur
mesh_terms: D001412:Bacillus subtilis / Q000201:enzymology / Q000235:genetics*; D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism; D002473:Cell Wall / Q000201:enzymology / Q000235:genetics; D003546:Cysteine Endopeptidases / Q000235:genetics; D003960:Diaminopimelic Acid / Q000378:metabolism; D024881:Drug Resistance, Bacterial / Q000235:genetics; D013293:Enterococcus faecalis / Q000201:enzymology / Q000235:genetics*; D009842:Oligopeptides / Q000378:metabolism; D046915:Penicillin-Binding Proteins / Q000235:genetics* / Q000378:metabolism; D010457:Peptidoglycan / Q000096:biosynthesis / Q000235:genetics*; D010458:Peptidyl Transferases / Q000235:genetics*; D017386:Sequence Homology, Amino Acid; D013045:Species Specificity; D013379:Substrate Specificity
chemical_list: D001426:Bacterial Proteins; D009842:Oligopeptides; D046915:Penicillin-Binding Proteins; D010457:Peptidoglycan; D003960:Diaminopimelic Acid; D010458:Peptidyl Transferases; D003546:Cysteine Endopeptidases