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

Abstract

Peptide deformylase (PDF) inhibitors have a strong potential to be used as a new class of antibiotics. However, recent studies have shown that the mitochondria of most eukaryotes, including humans, contain an essential PDF, PDF1A. The crystal structure of the Arabidopsis thaliana PDF1A (AtPDF1A), considered representative of PDF1As in general, has been determined. This structure displays several similarities to that of known bacterial PDFs. AtPDF1A behaves as a dimer, with the C-terminal residues responsible for linking the two subunits. This arrangement is similar to that of Leptospira interrogans PDF, the only other dimeric PDF identified to date. AtPDF1A is the first PDF for which zinc has been identified as the catalytic ion. However, the zinc binding pocket does not differ from the binding pockets of PDFs with iron rather than zinc. The crystal structure of AtPDF1A in complex with a substrate analog revealed that the substrate binding pocket of PDF1A displays strong modifications. The S1 binding pocket is significantly narrower, due to the creation of a floor from residues present in all PDF1As but not in bacterial PDFs. A true S3 pocket is created by the residues of a helical CD-loop, which is very long in PDF1As. Finally, these modified substrate binding pockets modify the position of the substrate in the active site. These differences provide guidelines for the design of bacterial PDF inhibitors that will not target mitochondrial PDFs.

Genome-Wide Identification and in Silico Analysis of Poplar Peptide Deformylases

“…In Arabidopsis, the two PDF1s form two subclasses: PDF1A that localizes to the mitochondria, and PDF1B that localizes to plastids [1,12]. These crystal structures of Arabidopsis PDF1A and PDF1B have been determined, not only indicating several similarities to bacterial PDFs and their function activity for the removal of the N -formyl group, but also showing several clear differences between AtPDF1A (At1g15390.1) and AtPDF1B (At5g14660.1) [8,9]. …”

“…Most PDFs are monomeric hydrolases and all contain three signature sequence motifs, comprising the active pocket of the enzyme and a metal cation: (i) G ψ G ψ AAXQ (motif 1); (ii) EGCLS (motif 2) and (iii) HE ψ DH (motif 3), where ψ is a hydrophobic amino acid [8,9]. The Cys of motif 2 and the two His residues of motif 3 stabilize metal ion coordination at the active site of PDF [8,9]. PDFs are important for some biological processes such as development of chloroplast in rice [10], and cell proliferation in humans [11].…”

“…Recent studies, together with the release of complete genome sequences for different organisms, have led to the identification of PDFs in eukaryotes; two PDFs have been identified in Arabidopsis [8,9], three in rice [10] and one in humans [11]. Since these PDFs do not contain the two insertions typical of PDF2 molecules, all eukaryotic PDFs are grouped as type 1 (PDF1).…”

“…Although amount of research efforts have been employed in exploring PDFs structure and function for several plant species, such as Arabidopsis [1,8,9,12] and rice [10], such research has not yet been directed towards woody trees. In order to identify all genes encoding PDFs and explore their function in poplar, we initiate one genome-wide investigation combined with in silico simulations.…”

Characterization of peptide deformylase homologues from Staphylococcus epidermidis

“…PDF enzymes from different species, including E. coli, S. aureus and others, have been cloned and both their threedimensional structures and co-crystal structures with a binding inhibitor have been reported (Baldwin et al, 2002;Cai et al, 2006;Chan et al, 1997;Fieulaine et al, 2005;Guilloteau et al, 2002;Robien et al, 2004;Smith et al, 2003;Zhou et al, 2005;Escobar-Alvarez et al, 2009). Although it has been reported recently that in some species such as Borrelia burgdorferi, Leptospira interrogans and Arabidopsis thaliana, the native PDFs exist as Zn 2+ -containing forms (Li et al, 2002;Nguyen et al, 2007), most native PDF enzymes in bacteria are unique Fe 2+ -containing metallohydrolases, which are highly active but unstable in vitro due to oxidation (Groche et al, 1998;Ragusa et al, 1998;Rajagopalan et al, 1997 (Chan et al, 1997;Meinnel et al, 1995Meinnel et al, , 1996, Ni 2+ (Groche et al, 1998;Ragusa et al, 1998;Wang et al, 2006) or Co 2+ (Rajagopalan et al, 2000;Dong & Liu, 2008), the enzymes retained their activities but were more stable, permitting further biochemical characterization and X-ray crystallographic analysis.…”