Structure of Peptide Deformylase and Identification of the Substrate Binding Site

Becker, Andreas; Schlichting, Ilme; Kabsch, Wolfgang; Schultz, Sabine; Wagner, A.

doi:10.1074/jbc.273.19.11413

Cited by 97 publications

(80 citation statements)

References 23 publications

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“…1). Substitutions at either of the analogous residues of E. coli PDF dramatically impair enzyme activity (3,4,8,18,20). The S. aureus defA gene, which also contains two substitutions in this motif ( Fig.…”

Section: Resultsmentioning

confidence: 99%

Resistance of Streptococcus pneumoniae to Deformylase Inhibitors Is Due to Mutations in defB

Margolis¹,

Hackbarth²,

López³

et al. 2001

Antimicrob Agents Chemother

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Resistance to peptide deformylase inhibitors in Escherichia coli or Staphylococcus aureus is due to inactivation of transformylase activity. Knockout experiments in Streptococcus pneumoniae R6x indicate that the transformylase (fmt) and deformylase (defB) genes are essential and that a def paralog (defA) is not. Actinoninresistant mutants of S. pneumoniae ATCC 49619 harbor mutations in defB but not in fmt. Reintroduction of the mutated defB gene into wild-type S. pneumoniae R6x recreates the resistance phenotype. The altered enzyme displays decreased sensitivity to actinonin.

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Section: Resultsmentioning

confidence: 99%

Resistance of Streptococcus pneumoniae to Deformylase Inhibitors Is Due to Mutations in defB

Margolis¹,

Hackbarth²,

López³

et al. 2001

Antimicrob Agents Chemother

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“…PEG molecules are known to occupy clefts that naturally bind elongated polymers such as either polypeptides in chaperones (55,56), in peptide deformylase (57,58), and in the neuronal calcium sensor (59) or long acyl chains in enzymes (59 -61) and in a periplasmic lipoprotein localization factor (62). The interaction network described for these examples is similar to the one observed in the P1D structure.…”

Section: Resultsmentioning

confidence: 99%

The Crystal Structure of the Periplasmic Domain of the Escherichia coli Membrane Protein Insertase YidC Contains a Substrate Binding Cleft

Ravaud

Stjepanović

Wild

et al. 2008

Journal of Biological Chemistry

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In bacteria the biogenesis of inner membrane proteins requires targeting and insertion factors such as the signal recognition particle and the Sec translocon. YidC is an essential membrane protein involved in the insertion of inner membrane proteins together with the Sec translocon, but also as a separate entity. YidC of Escherichia coli is a member of the conserved YidC (in bacteria)/Oxa1 (in mitochondria)/Alb3 (in chloroplasts) protein family and contains six transmembrane segments and a large periplasmic domain (P1). We determined the crystal structure of the periplasmic domain of YidC from E. coli (P1D) at 1.8 Å resolution. The structure of P1D shows the conserved ␤-supersandwich fold of carbohydrate-binding proteins and an ␣-helical linker region at the C terminus that packs against the ␤-supersandwich by a highly conserved interface. P1D exhibits an elongated cleft of similar architecture as found in the structural homologs. However, the electrostatic properties and molecular details of the cleft make it unlikely to interact with carbohydrate substrates. The cleft in P1D is occupied by a polyethylene glycol molecule suggesting an elongated peptide or acyl chain as a natural ligand. The region of P1D previously reported to interact with SecF maps to a surface area in the vicinity of the cleft. The conserved C-terminal region of the P1 domain was reported to be essential for the membrane insertase function of YidC. The analysis of this region suggests a role in membrane interaction and/or in the regulation of YidC interaction with binding partners.

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“…1), diagnostic of peptide deformylases (9,18,24). It appears that DefA is missing consensus cysteine and histidine residues that have been shown in the E. coli enzyme to bind to a metal cofactor essential for deformylase activity (3,4,(22)(23)(24)30). The defA gene was therefore considered unlikely to encode a functional peptide deformylase.…”

Section: Identification Of S Aureus Def Homologsmentioning

confidence: 99%

Peptide Deformylase in Staphylococcus aureus : Resistance to Inhibition Is Mediated by Mutations in the Formyltransferase Gene

Margolis¹,

Hackbarth²,

Young³

et al. 2000

Antimicrob Agents Chemother

123

115

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Peptide deformylase, a bacterial enzyme, represents a novel target for antibiotic discovery. Two deformylase homologs, defA and defB, were identified in Staphylococcus aureus. The defA homolog, located upstream of the transformylase gene, was identified by genomic analysis and was cloned from chromosomal DNA by PCR. A distinct homolog, defB, was cloned from an S. aureus genomic library by complementation of the arabinosedependent phenotype of a P BAD -def Escherichia coli strain grown under arabinose-limiting conditions. Overexpression in E. coli of defB, but not defA, correlated to increased deformylase activity and decreased susceptibility to actinonin, a deformylase-specific inhibitor. The defB gene could not be disrupted in wild-type S. aureus, suggesting that this gene, which encodes a functional deformylase, is essential. In contrast, the defA gene could be inactivated; the function of this gene is unknown. Actinonin-resistant mutants grew slowly in vitro and did not show cross-resistance to other classes of antibiotics. When compared to the parent, an actinonin-resistant strain produced an attenuated infection in a murine abscess model, indicating that this strain also has a growth disadvantage in vivo. Sequence analysis of the actinonin-resistant mutants revealed that each harbors a loss-of-function mutation in the fmt gene. Susceptibility to actinonin was restored when the wild-type fmt gene was introduced into these mutant strains. An S. aureus ⌬fmt strain was also resistant to actinonin, suggesting that a functional deformylase activity is not required in a strain that lacks formyltransferase activity. Accordingly, the defB gene could be disrupted in an fmt mutant.

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Structure of Peptide Deformylase and Identification of the Substrate Binding Site

Cited by 97 publications

References 23 publications

Resistance of Streptococcus pneumoniae to Deformylase Inhibitors Is Due to Mutations in defB

Resistance of Streptococcus pneumoniae to Deformylase Inhibitors Is Due to Mutations in defB

The Crystal Structure of the Periplasmic Domain of the Escherichia coli Membrane Protein Insertase YidC Contains a Substrate Binding Cleft

Peptide Deformylase in Staphylococcus aureus : Resistance to Inhibition Is Mediated by Mutations in the Formyltransferase Gene

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