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Epimerization and substrate gating by a TE domain in β-lactam antibiotic biosynthesis
Nicole M Gaudelli1, Craig A Townsend1
1Department of Chemistry, Johns Hopkins University, Baltimore, Maryland, USA.
Nature Chemical Biology
|February 18, 2014
Summary
This study reveals a novel dual-function thioesterase (TE) domain in nocardicin A biosynthesis. This TE domain acts as a gatekeeper, catalyzing epimerization and releasing the pentapeptide β-lactam precursor, nocardicin G.
Area of Science:
- Biochemistry
- Natural Product Biosynthesis
- Enzymology
Background:
- Nonribosomal peptide synthetases (NRPS) synthesize bioactive natural products via a thiotemplate mechanism.
- C-terminal thioesterase (TE) domains typically release products through hydrolysis or macrocyclization.
Purpose of the Study:
- To investigate the unprecedented dual-function TE domain in nocardicin A biosynthesis.
- To elucidate the mechanism of nocardicin G formation, the precursor to nocardicins.
Main Methods:
- Biochemical characterization of the nocardicin TE domain.
- Kinetic analysis using stereodefined peptide substrates.
- Investigation of substrate discrimination and catalytic functions.
Main Results:
- The nocardicin TE domain showed stringent discrimination against peptide substrates, failing to hydrolyze them.
- Prior monocyclic β-lactam formation at an L-seryl site was necessary to overcome substrate discrimination.
- The TE domain functions as a gatekeeper, holding the peptide until β-lactam formation, then catalyzing epimerization and thioesterase cleavage.
- This process releases nocardicin G, a pentapeptide β-lactam.
Conclusions:
- The nocardicin TE domain possesses a novel dual function: gatekeeping and catalyzing epimerization, in addition to thioesterase cleavage.
- This unique mechanism is crucial for the biosynthesis of nocardicin G, the precursor to monocyclic β-lactam antibiotics.
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