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Testing the Role of Multicopy Plasmids in the Evolution of Antibiotic Resistance
Published on: May 2, 2018
Structural and functional insights into esterase-mediated macrolide resistance
Michał Zieliński1,2, Jaeok Park1,3, Barry Sleno1,2
1Department of Biochemistry, McGill University, Montréal, QC, Canada.
Abstract:
Macrolides are a class of antibiotics widely used in both medicine and agriculture. Unsurprisingly, as a consequence of their exensive usage a plethora of resistance mechanisms have been encountered in pathogenic bacteria. One of these resistance mechanisms entails the enzymatic cleavage of the macrolides' macrolactone ring by erythromycin esterases (Eres). The most frequently identified Ere enzyme is EreA, which confers resistance to the majority of clinically used macrolides. Despite the role Eres play in macrolide resistance, research into this family enzymes has been sparse. Here, we report the first three-dimensional structures of an erythromycin esterase, EreC. EreC is an extremely close homologue of EreA, displaying more than 90% sequence identity. Two structures of this enzyme, in conjunction with in silico flexible docking studies and previously reported mutagenesis data allowed for the proposal of a detailed catalytic mechanism for the Ere family of enzymes, labeling them as metal-independent hydrolases. Also presented are substrate spectrum assays for different members of the Ere family. The results from these assays together with an examination of residue conservation for the macrolide binding site in Eres, suggests two distinct active site archetypes within the Ere enzyme family.
Insights
Erythromycin esterases (Eres) confer macrolide antibiotic resistance by cleaving the macrolactone ring. This study reveals the structures and catalytic mechanism of EreC, identifying distinct active site archetypes within the Ere enzyme family.
Area of Science:
- Microbiology
- Structural Biology
- Biochemistry
Background:
- Macrolide antibiotics are extensively used in medicine and agriculture.
- Macrolide resistance in bacteria is a growing concern, often mediated by enzymes.
- Erythromycin esterases (Eres) are key enzymes conferring resistance through macrolactone ring cleavage.
Purpose of the Study:
- To elucidate the structural basis of macrolide resistance conferred by erythromycin esterases (Eres).
- To determine the catalytic mechanism of the Ere enzyme family.
- To investigate the diversity of active site structures within the Ere family.
Main Methods:
- X-ray crystallography to determine the 3D structures of EreC.
- In silico flexible docking studies.
- Substrate spectrum assays and analysis of residue conservation.
Main Results:
- Two distinct 3D structures of EreC, a close homolog of EreA, were determined.
- A detailed catalytic mechanism was proposed, classifying Eres as metal-independent hydrolases.
- Substrate assays and structural analysis revealed two distinct active site archetypes within the Ere family.
Conclusions:
- The structural and mechanistic insights into EreC provide a foundation for understanding Ere-mediated macrolide resistance.
- The identification of distinct active site archetypes suggests potential for developing targeted inhibitors.
- Further research into the Ere enzyme family is warranted to combat macrolide resistance.
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