Assays to Detect and Characterize Synthetic Agents that Inhibit the ErmC Methyltransferase.

Clancy, Joanna; Schmieder, Brenda J.; Petitpas, Joan W.; Manousos, Mary; Williams, John A.; Faiella, J.; Girard, Arthur E.; McGuirk, Paul R.

doi:10.7164/antibiotics.48.1273

Cited by 32 publications

(33 citation statements)

References 13 publications

Supporting

Mentioning

Contrasting

Unclassified

Order By: Relevance

“…[16] The residues that line up the cofactor binding site are conserved among different MTase families, while the substrate base binding site differs depending on the type of substrate and the type of methylation catalyzed. [17,18] For instance, nearly all MTases that methylate exocyclic amino groups in RNA or DNA to yield m 6 A, m 2 G, or m 4 C, exhibit a characteristic motif: (N/D/S)-(I/P)-P-(Y/F/W/H). [19] In the protein-substrate co-crystal structure of DNA:m 6 A-MTase M.TaqI, a relatively close homolog of Erm MTases, the aromatic residue at the terminal position stabilizes the methylation substrate base by face-to-face stacking interactions, while the main-chain carboxyl group of the conserved Pro residue and the side chain of the semi-conserved Asn hydrogen bond the target NH 2 group and stabilize it with respect to the methyl group donor.…”

Section: Identification Of the Optimal Receptor Conformationmentioning

confidence: 99%

“…[17,18] For instance, nearly all MTases that methylate exocyclic amino groups in RNA or DNA to yield m 6 A, m 2 G, or m 4 C, exhibit a characteristic motif: (N/D/S)-(I/P)-P-(Y/F/W/H). [19] In the protein-substrate co-crystal structure of DNA:m 6 A-MTase M.TaqI, a relatively close homolog of Erm MTases, the aromatic residue at the terminal position stabilizes the methylation substrate base by face-to-face stacking interactions, while the main-chain carboxyl group of the conserved Pro residue and the side chain of the semi-conserved Asn hydrogen bond the target NH 2 group and stabilize it with respect to the methyl group donor. [20] Mutagenesis of the corresponding residues in ErmC' has shown the aromatic Tyr 104 residue to be absolutely essential for ErmC' activity, while the semi-conserved Asn 101 was found to be important but not absolutely essential.…”

Section: Identification Of the Optimal Receptor Conformationmentioning

confidence: 99%

“…A highthroughput screening analysis has detected compounds that selectively inhibit the ErmC' enzyme in the nano-to micromolar range in vitro, but which failed, however, to inhibit macrolide resistance in an in vivo mouse model. [6] Although these synthetic leads were regarded as a promising starting point for the design of more potent inhibitors, without the knowledge of the protein binding mode it has been difficult to optimize their activity. To our knowledge, no follow-up study has appeared since the initial report, either as a patent or publication.…”

Section: Introductionmentioning

confidence: 99%

“…The resistance to all these antibiotics is caused by mono-or dimethylation of 23S rRNA at the N6 position of adenosine 2058 (E. coli numbering), which is a pivotal nucleoside for the binding of MLS antibiotics. The methylation of A2058 to m 6 A and subsequently to m 6 2 A is introduced by enzymes encoded by a group of genes termed erm (erythromycin resistance methyltransferases). [2] Horizontal transfer of erm genes to numerous pathogenic strains and global dissemination of resistant clones and their descendants are major components of the present-day macrolide resistance…”

Section: Introductionmentioning

confidence: 99%

See 3 more Smart Citations

Virtual Screening and Experimental Verification to Identify Potential Inhibitors of the ErmC Methyltransferase Responsible for Bacterial Resistance against Macrolide Antibiotics

et al. 2008

View full text Add to dashboard Cite

Methyltransferases from the Erm family catalyze S-adenosyl-L-methionine-dependent modification of a specific adenine residue in bacterial 23S rRNA, thereby conferring resistance to clinically important macrolide, lincosamide, and streptogramin B antibiotics. Thus far, no inhibitors of these enzymes have been identified or designed that would effectively abolish the resistance in vivo. We used the crystal structure of ErmC' methyltransferase as a target for structure-based virtual screening of a database composed of 58,679 lead-like compounds. Among 77 compounds selected for experimental validation (63 predicted to bind to the catalytic pocket and 14 compounds predicted to bind to the putative RNA binding site), we found several novel inhibitors that decrease the minimal inhibitory concentration of a macrolide antibiotic erythromycin toward an Escherichia coli strain that constitutively expresses ErmC'. Eight of them have IC(50) values in the micromolar range. Analysis of docking models of the identified inhibitors suggests a novel strategy to develop potent and clinically useful inhibitors.

show abstract

Section: Identification Of the Optimal Receptor Conformationmentioning

confidence: 99%

Section: Identification Of the Optimal Receptor Conformationmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

Virtual Screening and Experimental Verification to Identify Potential Inhibitors of the ErmC Methyltransferase Responsible for Bacterial Resistance against Macrolide Antibiotics

et al. 2008

View full text Add to dashboard Cite

show abstract

“…Attempts have been made to discover Erm methyltransferase inhibitors that maintain the effectiveness of macrolide antibiotics in the face of the increasing frequency of resistant isolates (2,3). The latter work (3) was part of a series of major structural studies of Erm methyltransferases (1,5,9).…”

mentioning

confidence: 99%

Modulation of Erm Methyltransferase Activity by Peptides Derived from Phage Display

Giannattasio

Weisblum

2000

Antimicrob Agents Chemother

View full text Add to dashboard Cite

Combinatorial peptide display on phage M13 protein pIII was used to discover peptide sequences that selectively bind to ErmC methyltransferase from Bacillus subtilis. One peptide, Ac-LSGVIAT-NH 2 , inhibited methylation in vitro with a 50% inhibitory concentration of 20 M. Interestingly, the set of six peptides which inhibited ErmC stimulated ErmSF, a homologous methyltransferase from Streptomyces fradiae. Thus, Ac-LSGVIAT-NH 2 may not act directly at the catalytic center of ErmC, but may modulate its activity by binding at a structurally unrelated, but functionally linked, site.The ErmC N-methyltransferase of Staphylococcus aureus and its close relative ErmCЈ from Bacillus subtilis confer resistance to erythromycin and related macrolide antibiotics. These enzymes act by specifically methylating a single adenine residue in the peptidyl transferase center of bacterial 23S rRNA (for reviews, see references 7 and 8). The same methylation confers coresistance to structurally unrelated lincosamide and streptogramin type B antibiotics. The three groups are collectively known as the macrolide-lincosamide-streptogramin B (MLS) antibiotics, and the form of resistance based on the Erm group of methyltransferases is found in a wide range of pathogens. The three-dimensional structures of the ErmAM and ErmCЈ methyltransferases have recently been solved by nuclear magnetic resonance (3, 9) and X-ray crystallography (1, 5), respectively. These developments bring us closer to rationally devising ligands that will selectively bind to Erm enzymes and possibly reduce the efficiency with which they confer MLS resistance.Macrolide antibiotics have served as a mainstay of antimicrobial therapy for approximately the last half century, especially in instances where the recipient of the antibiotic was allergic to beta-lactam antibiotics. Attempts have been made to discover Erm methyltransferase inhibitors that maintain the effectiveness of macrolide antibiotics in the face of the increasing frequency of resistant isolates (2, 3). The latter work (3) was part of a series of major structural studies of Erm methyltransferases (1, 5, 9).Nonpeptide ligands which displace an inhibitory peptide from its binding site on the enzyme might, themselves, have inhibitory activity. The use of inhibitory peptides might thus serve as a platform for the discovery of nonpeptide inhibitors of Erm enzymes. A proposed way to achieve this would be based on the discovery of test ligands to displace an inhibitory peptide from its association with its cognate Erm target. The displacement of an inhibitory ligand would also be easier to measure than methyltransferase activity.Large-scale screening of inhibitory ligands by a direct assay of methyltransferase catalytic activity is cumbersome since it requires the separation of product (methyl-labeled 23S rRNA) from substrate (unreacted S-adenosyl-L-methionine). We report the use of combinatorial phage display to discover peptides that inhibit ErmCЈ methylase activity and which might serve as displaceable ligands in ...

show abstract

Kohlenhydrate in der Antibiotikaforschung: ein neuer Ansatz zur Resistenzbekämpfung

Ritter

Wong

2001

Angew. Chem.

View full text Add to dashboard Cite

Assays to Detect and Characterize Synthetic Agents that Inhibit the ErmC Methyltransferase.

Cited by 32 publications

References 13 publications

Virtual Screening and Experimental Verification to Identify Potential Inhibitors of the ErmC Methyltransferase Responsible for Bacterial Resistance against Macrolide Antibiotics

Virtual Screening and Experimental Verification to Identify Potential Inhibitors of the ErmC Methyltransferase Responsible for Bacterial Resistance against Macrolide Antibiotics

Modulation of Erm Methyltransferase Activity by Peptides Derived from Phage Display

Kohlenhydrate in der Antibiotikaforschung: ein neuer Ansatz zur Resistenzbekämpfung

Contact Info

Product

Resources

About