Novel Mechanism of Resistance to Glycopeptide Antibiotics in Enterococcus faecium

Abstract

Glycopeptides and ␤-lactams are the major antibiotics available for the treatment of infections due to Gram-positive bacteria. Emergence of cross-resistance to these drugs by a single mechanism has been considered as unlikely because they inhibit peptidoglycan polymerization by different mechanisms. Therapeutic usage of antibiotics has invariably led to emergence of resistance in target pathogens and members of commensal flora submitted to the same selective pressure. This scenario differs greatly between antibiotics with respect to the time period separating initial drug marketing and emergence of resistance (1). Detoxification of penicillin was recognized before the first therapeutic usage of the drug (2), and acquisition of resistance was immediately a medical concern (3). In contrast, acquired resistance to glycopeptides was detected for the first time in enterococci 28 years after vancomycin had been launched in 1956 (4). During this unusually long delay, glycopeptides were considered as "irresistible" antibiotics, and it has been rationalized that this class of drugs remained unharmed because of its unique mode of action (5). The drugs bind to the peptidyl-D-Ala 4 -D-Ala 5 extremity of peptidoglycan precursors at the bacterial cell surface and block the transglycosylation and transpeptidation reactions by steric hindrance (Fig. 1A). This mode of action implies that acquisition of resistance cannot be easily achieved because modification of the target is limited by the specificity of multiple biosynthetic enzymes (5), including the Ddl ligase for synthesis of D-Ala-D-Ala, the MurF ligase for addition of the resulting dipeptide to the nascent cytoplasmic precursor, and the D,D-transpeptidases for peptidoglycan crosslinking in the last step of peptidoglycan polymerization (6). The latter reaction is catalyzed by penicillin-binding proteins (PBPs) 2 that cleave the D-Ala 4 -D-Ala 5 bond of a donor stem pentapeptide and link the carboxyl of D-Ala 4 to the amino group of the side chain carried by the third residue of an acceptor stem peptide (Fig. 1A).Glycopeptide resistance has emerged in the enterococci (4) by acquisition of transposon Tn1546, which mediates production of precursors terminating in D-lactate (D-Lac) instead of D-Ala (7,8). The substitution leads to a 1,000-fold reduction in the affinity of vancomycin for its target (9).

The Peptidoglycan of Stationary-Phase Mycobacterium tuberculosis Predominantly Contains Cross-Links Generated by l,d -Transpeptidation

“…D-Ala, mostly present at the fourth position, was replaced by Asn, presumably of the D configuration, in a minority of the stem peptides. Since the latter amino acid was abundant in the culture me- dium, its presence in muropeptides could result from the exchange of D-Ala with D-Asn in the peptidoglycan due to an L,D-transpeptidation reaction, as previously discussed for E. faecium (9,18).…”

Specificity of L,D-Transpeptidases from Gram-positive Bacteria Producing Different Peptidoglycan Chemotypes

“…Conclusions-Bypass of the D,D-transpeptidase activity of the PBPs by the L,D-transpeptidase activity of Ldt fm confers high level cross-resistance to ␤-lactams and glycopeptides in E. faecium because these drugs do not interact with the enzyme and its substrate, respectively (9). Ldt fm is the first functionally characterized member of a novel family of cysteine peptidases that are widespread in both Gram-positive and Gram-negative bacteria (18,35).…”

“…Strikingly, production of D-lactate-ending precursors can also have an impact on the activity of ␤-lactams in the enterococci and staphylococci, presumably because low affinity PBPs responsible for ␤-lactam resistance cannot function with modified precursors (6,8). Total elimination of D-Ala 5 by hydrolysis of the C-terminal residue of pentapeptide stems is an alternative mechanism of glycopeptide resistance in mutants of Enterococcus faecium selected in laboratory conditions (9). Because PBPs cannot function with tetrapeptide donors, peptidoglycan cross-linking in these mutants requires an L,Dtranspeptidase (Ldt fm ) that cleaves the L-Lys 3 -D-Ala 4 peptide bond of the donor and links the carboxyl of L-Lys 3 to the side chain amine of the acceptor (Fig.…”

Unexpected Inhibition of Peptidoglycan LD-Transpeptidase from Enterococcus faecium by the β-Lactam Imipenem

“…Ldt fm exclusively uses donor substrate carrying a tetrapeptide stem (16 (14,16). Full elimination of pentapeptide stems ending in D-Ala 5 by this DDcarboxypeptidase leads to high level cross-resistance to a second family of antibiotics, the glycopeptides, that bind to the peptidyl-D-Ala 4 -D-Ala 5 extremity of peptidoglycan precursors (19).…”