Dataset Information

Mycobacterium abscessus l,d-Transpeptidases Are Susceptible to Inactivation by Carbapenems and Cephalosporins but Not Penicillins.

ABSTRACT: As a growing number of clinical isolates of Mycobacterium abscessus are resistant to most antibiotics, new treatment options that are effective against these drug-resistant strains are desperately needed. The majority of the linkages in the cell wall peptidoglycan of M. abscessus are synthesized by nonclassical transpeptidases, namely, the l,d-transpeptidases. Emerging evidence suggests that these enzymes represent a new molecular vulnerability in this pathogen. Recent studies have demonstrated that inhibition of these enzymes by the carbapenem class of ?-lactams determines their activity against Mycobacterium tuberculosis Here, we studied the interactions of ?-lactams with two l,d-transpeptidases in M. abscessus, namely, LdtMab1 and LdtMab2, and found that both the carbapenem and cephalosporin, but not penicillin, subclasses of ?-lactams inhibit these enzymes. Contrary to the commonly held belief that combination therapy with ?-lactams is redundant, doripenem and cefdinir exhibit synergy against both pansusceptible M. abscessus and clinical isolates that are resistant to most antibiotics, which suggests that dual-?-lactam therapy has potential for the treatment of M. abscessus Finally, we solved the first crystal structure of an M. abscessus l,d-transpeptidase, LdtMab2, and using substitutions of critical amino acids in the catalytic site and computational simulations, we describe the key molecular interactions between this enzyme and ?-lactams, which provide an insight into the molecular basis for the relative efficacy of different ?-lactams against M. abscessus.

SUBMITTER: Kumar P

PROVIDER: S-EPMC5610527 | biostudies-literature | 2017 Oct

REPOSITORIES: biostudies-literature

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Mycobacterium abscessus l,d-Transpeptidases Are Susceptible to Inactivation by Carbapenems and Cephalosporins but Not Penicillins.

Kumar Pankaj P Chauhan Varsha V Silva José Rogério A JRA Lameira Jerônimo J d'Andrea Felipe B FB Li Shao-Gang SG Ginell Stephan L SL Freundlich Joel S JS Alves Cláudio Nahum CN Bailey Scott S Cohen Keira A KA Lamichhane Gyanu G

Antimicrobial agents and chemotherapy 20170922 10

As a growing number of clinical isolates of <i>Mycobacterium abscessus</i> are resistant to most antibiotics, new treatment options that are effective against these drug-resistant strains are desperately needed. The majority of the linkages in the cell wall peptidoglycan of <i>M. abscessus</i> are synthesized by nonclassical transpeptidases, namely, the l,d-transpeptidases. Emerging evidence suggests that these enzymes represent a new molecular vulnerability in this pathogen. Recent studies have ...[more]

PMID: 28760902

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Project description:Mycobacterium abscessus is a highly drug-resistant nontuberculous mycobacterium (NTM). Efforts to discover new treatments for M. abscessus infections are accelerating, with a focus on cell wall synthesis proteins (M. abscessus l,d-transpeptidases 1 to 5 [LdtMab1 to LdtMab5] and d,d-carboxypeptidase) that are targeted by β-lactam antibiotics. A challenge to this approach is the presence of chromosomally encoded β-lactamase (BlaMab). Using a mechanism-based approach, we found that a novel ceftaroline-imipenem combination effectively lowered the MICs of M. abscessus isolates (MIC50 ≤ 0.25 μg/ml; MIC90 ≤ 0.5 μg/ml). Combining ceftaroline and imipenem with a β-lactamase inhibitor, i.e., relebactam or avibactam, demonstrated only a modest effect on susceptibility compared to each of the β-lactams alone. In steady-state kinetic assays, BlaMab exhibited a lower Ki app (0.30 ± 0.03 μM for avibactam and 136 ± 14 μM for relebactam) and a higher acylation rate for avibactam (k2/K = 3.4 × 105 ± 0.4 × 105 M-1 s-1 for avibactam and 6 × 102 ± 0.6 × 102 M-1 s-1 for relebactam). The kcat/Km was nearly 10-fold lower for ceftaroline fosamil (0.007 ± 0.001 μM-1 s-1) than for imipenem (0.056 ± 0.006 μM-1 s-1). Timed mass spectrometry captured complexes of avibactam and BlaMab, LdtMab1, LdtMab2, LdtMab4, and d,d-carboxypeptidase, whereas relebactam bound only BlaMab, LdtMab1, and LdtMab2 Interestingly, LdtMab1, LdtMab2, LdtMab4, LdtMab5, and d,d-carboxypeptidase bound only to imipenem when incubated with imipenem and ceftaroline fosamil. We next determined the binding constants of imipenem and ceftaroline fosamil for LdtMab1, LdtMab2, LdtMab4, and LdtMab5 and showed that imipenem bound >100-fold more avidly than ceftaroline fosamil to LdtMab1 and LdtMab2 (e.g., Ki app or Km of LdtMab1 = 0.01 ± 0.01 μM for imipenem versus 0.73 ± 0.08 μM for ceftaroline fosamil). Molecular modeling indicates that LdtMab2 readily accommodates imipenem, but the active site must widen to ≥8 Å for ceftaroline to enter. Our analysis demonstrates that ceftaroline and imipenem binding to multiple targets (l,d-transpeptidases and d,d-carboxypeptidase) and provides a mechanistic rationale for the effectiveness of this dual β-lactam combination in M. abscessus infections.

Dataset Information

Mycobacterium abscessus l,d-Transpeptidases Are Susceptible to Inactivation by Carbapenems and Cephalosporins but Not Penicillins.

Publications

Mycobacterium abscessus l,d-Transpeptidases Are Susceptible to Inactivation by Carbapenems and Cephalosporins but Not Penicillins.

Similar Datasets

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