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A Protocol for Functional Assessment of Whole-Protein Saturation Mutagenesis Libraries Utilizing High-Throughput Sequencing
Published on: July 3, 2016
Structural aspects for evolution of beta-lactamases from penicillin-binding proteins
Samy O Meroueh1, George Minasov, Wenlin Lee
1Department of Chemistry and Biochemistry, University of Notre Dame, Notre Dame, Indiana 46556, USA.
Journal of the American Chemical Society
|August 9, 2003
Summary
Bacterial resistance enzymes, beta-lactamases, evolved distinct active sites to prevent interaction with peptidoglycan, unlike penicillin-binding proteins. Structural analysis and simulations confirm modifications that abolish peptidoglycan binding, enabling antibiotic resistance.
Area of Science:
- Biochemistry
- Structural Biology
- Microbiology
Background:
- Penicillin-binding proteins (PBPs) and beta-lactamases share evolutionary origins.
- PBPs synthesize bacterial cell walls using peptidoglycan.
- Beta-lactamases confer resistance to beta-lactam antibiotics.
Purpose of the Study:
- To investigate the structural modifications enabling beta-lactamases to function as antibiotic resistance enzymes.
- To test the hypothesis that beta-lactamases evolved to avoid interaction with peptidoglycan, the substrate for PBPs.
Main Methods:
- Synthesis of a cephalosporin analogue (compound 6).
- X-ray crystallography to determine the structure of the cephalosporin-AmpC beta-lactamase complex.
- Molecular dynamics simulation of the complex.
Main Results:
- The X-ray structure revealed the absence of a peptidoglycan interaction surface in the beta-lactamase active site.
- A peptide insertion in the beta-lactamase active site prevents interaction with peptidoglycan strands.
- Molecular dynamics simulations showed the bound ligand is not stabilized, indicating a lack of specific binding.
Conclusions:
- Structural modifications in beta-lactamases prevent interaction with peptidoglycan.
- These changes are crucial for the enzyme's catalytic proficiency in antibiotic resistance.
- The study elucidates the evolutionary divergence of beta-lactamases from PBPs.
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