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Mechanism of the C5 Stereoinversion Reaction in the Biosynthesis of Carbapenem Antibiotics

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Science  07 Mar 2014:
Vol. 343, Issue 6175, pp. 1140-1144
DOI: 10.1126/science.1248000

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Carbapenems Through the Looking Glass

The carbapenem class of antibiotics is a critical weapon in the ongoing fight against drug-resistant bacteria. Microbial biosynthesis of these compounds, which contain a strained β-lactam ring motif, proceeds via a precursor that has the wrong configuration at one of the ring carbons. Chang et al. (p. 1140) combined x-ray crystallography with multiple spectroscopic probes to map out the mechanism by which the CarC enzyme inverts the precursor configuration to its mirror image.

Abstract

The bicyclic β-lactam/2-pyrrolidine precursor to all carbapenem antibiotics is biosynthesized by attachment of a carboxymethylene unit to C5 of l-proline followed by β-lactam ring closure. Carbapenem synthase (CarC), an Fe(II) and 2-(oxo)glutarate (Fe/2OG)–dependent oxygenase, then inverts the C5 configuration. Here we report the structure of CarC in complex with its substrate and biophysical dissection of its reaction to reveal the stereoinversion mechanism. An Fe(IV)-oxo intermediate abstracts the hydrogen (H•) from C5, and tyrosine 165, a residue not visualized in the published structures of CarC lacking bound substrate, donates H• to the opposite face of the resultant radical. The reaction oxidizes the Fe(II) cofactor to Fe(III), limiting wild-type CarC to one turnover, but substitution of the H•-donating tyrosine disables stereoinversion and confers to CarC the capacity for catalytic substrate oxidation.

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