Peptide deformylase (PDF) is essential in prokaryotes and absent in mammalian cells, thus making it an attractive target for the discovery of novel antibiotics. We have identified actinonin, a naturally occurring antibacterial agent, as a potent PDF inhibitor. The dissociation constant for this compound was 0.3 x 10(-)(9) M against Ni-PDF from Escherichia coli; the PDF from Staphylococcus aureus gave a similar value. Microbiological evaluation revealed that actinonin is a bacteriostatic agent with activity against Gram-positive and fastidious Gram-negative microorganisms. The PDF gene, def, was placed under control of P(BAD) in E. coli tolC, permitting regulation of PDF expression levels in the cell by varying the external arabinose concentration. The susceptibility of this strain to actinonin increases with decreased levels of PDF expression, indicating that actinonin inhibits bacterial growth by targeting this enzyme. Actinonin provides an excellent starting point from which to derive a more potent PDF inhibitor that has a broader spectrum of antibacterial activity.

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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Oxazolidinones comprise an important class of antibacterial protein synthesis inhibitors. Myelosuppression and monoamine oxidase inhibition (MAOI) are key independent causes for limiting adverse effects in therapy with the sole approved drug of this class, linezolid. This annotation describes a novel oxazolidinone agent, (S)-5-((isoxazol-3-ylamino)methyl)-3-(2,3,5-trifluoro-4-(4-oxo-3,4-dihydropyridin-1(2H)-yl)phenyl)oxazolidin-2-one (MRX-I), distinguished by its high activity against Gram-positive pathogens coupled with markedly reduced potential for myelosuppression and MAOI. The medical need, medicinal chemistry rationale, preclinical data, and phase I clinical trial summary for this new agent are reviewed herein.