Abstract
The alpha-amino acid ester hydrolase (AEH) from Acetobacter turbidans is a bacterial enzyme catalyzing the hydrolysis and synthesis of beta-lactam antibiotics. The crystal structures of the native enzyme, both unliganded and in complex with the hydrolysis product D-phenylglycine are reported, as well as the structures of an inactive mutant (S205A) complexed with the substrate ampicillin, and an active site mutant (Y206A) with an increased tendency to catalyze antibiotic production rather than hydrolysis. The structure of the native enzyme shows an acyl binding pocket, in which D-phenylglycine binds, and an additional space that is large enough to accommodate the beta-lactam moiety of an antibiotic. In the S205A mutant, ampicillin binds in this pocket in a non-productive manner, making extensive contacts with the side chain of Tyr(112), which also participates in oxyanion hole formation. In the Y206A mutant, the Tyr(112) side chain has moved with its hydroxyl group toward the catalytic serine. Because this changes the properties of the beta-lactam binding site, this could explain the increased beta-lactam transferase activity of this mutant.
Highlights
Thirty years ago, several bacterial strains, such as Acetobacter turbidans and Xanthomonas citri, were identified that were able to efficiently produce semi-synthetic -lactam antibiotics from -lactam nuclei produced by fermentation, and synthetic acyl compounds with an ␣-amino group [1]
Native Structure: Oligomeric State—The structures of the WT, WT1⁄7D-phenylglycine, Y206A, and S205A acid ester hydrolase (AEH) were successfully determined by x-ray crystallography
Because the active sites are found on the inside of the sphere, this tetrameric oligomerization state would restrict the AEHs to converting small compounds
Summary
Several bacterial strains, such as Acetobacter turbidans and Xanthomonas citri, were identified that were able to efficiently produce semi-synthetic -lactam antibiotics from -lactam nuclei produced by fermentation, and synthetic acyl compounds with an ␣-amino group [1]. The backbone amide of Tyr206, which directly follows the catalytic serine in the sequence, and the phenolic OH of the Tyr112 side chain form an oxyanion hole similar to that observed in X. citri AEH, CocE, and PepX [7,8,9].
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