Structural Basis for the Design of Antibiotics Targeting Peptide Deformylase^,

Abstract

While protein synthesis in bacteria begins with a formylated methionine, the formyl group of the nascent polypeptide is removed by peptide deformylase. Since eukaryotic protein synthesis does not involve formylation and deformylation at the N-terminus, there has been increasing interest in peptide deformylase as a potential target for antibacterial chemotherapy. Toward this end and to aid in the design of effective antibiotics targeting peptide deformylase, the structures of the protein-inhibitor complexes of both the cobalt and the zinc containing Escherichia coli peptide deformylase bound to the transition-state analogue, (S)-2-O-(H-phosphonoxy)-L-caproyl-L-leucyl-p-nitroanilide (PCLNA), have been determined. The proteins for both deformylase-inhibitor complexes show basically the same fold as for the native enzyme. The PCLNA inhibitor adopts an extended conformation and fits nicely into a hydrophobic cavity located near the metal site. On the basis of these structures, guidelines for the design of high-affinity deformylase inhibitors are suggested. As our results show that the protein residues which interact with the PCLNA inhibitor are conserved over a wide variety of species, we suggest that antibiotics targeting deformylase could have wide applicability.

The Crystal Structure of Mitochondrial (Type 1A) Peptide Deformylase Provides Clear Guidelines for the Design of Inhibitors Specific for the Bacterial Forms

“…The active site cavity is also well conserved, especially in terms of metal chelation, and the reaction mechanism has been studied in detail. Based on the observed structural similarities, we suggest that the catalytic mechanism of eukaryotic PDFs is strictly identical to that of bacterial PDFs (13,18,19), although PDF1A is highly active as a zinc enzyme, and bacterial PDFs in which the iron is replaced by zinc are inactive.…”

“…6). Hence, in the complex made between EcPDF and a transition analog mimicking Met-Leu-para-nitroanilide, the para-nitroanilide moiety makes a lid burying the Met mimic side chain into a hydrophobic cleft (19). This effect participates in strongly increasing the binding constant of this compound.…”

“…However, these differences do not affect folding of the active site. We and others have reported the three-dimensional structures of PDFs from various bacteria, in complex with the catalytic metal cation and/or with a ligand, such as an inhibitor or a product of the deformylation reaction (17)(18)(19). No significant differences in overall or active site structure were observed between the two forms.…”

“…No significant differences in overall or active site structure were observed between the two forms. The determination of these structures led to identification of the metal and peptide binding modes and elucidation of a catalytic mechanism for the deformylation reaction (13,18,19). For 3 decades, it has been widely accepted that eukaryotes have no peptide deformylase (20,21).…”

“…Since PDF was also shown to be present in several human parasites, such as the causative agent of malaria, it was also considered a good target for antiparasitic agents (23). The structural comparison of several PDFs in complex with an inhibitor has provided guidelines for the design of high affinity PDF inhibitors (PDF-In) (16,19). Actinonin, a pseudotripeptide hydroxamate derivative, has been shown to target peptide deformylases specifically and therefore to have natural antibiotic activity (24,25).…”