Discovery of 4-Pyridone Derivatives as Specific Inhibitors of Enoyl-Acyl Carrier Protein Reductase (FabI) with Antibacterial Activity against Staphylococcus aureus

ABSTRACT The MICs of triclosan for 31 clinical isolates of Staphylococcus aureus were 0.016 μg/ml (24 strains), 1 to 2 μg/ml (6 strains), and 0.25 μg/ml (1 strain). All the strains for which triclosan MICs were elevated (>0.016 μg/ml) showed three- to fivefold increases in their levels of enoyl-acyl carrier protein (ACP) reductase (FabI) production. Furthermore, strains for which triclosan MICs were 1 to 2 μg/ml overexpressed FabI with an F204C alteration. Binding studies with radiolabeled NAD+ demonstrated that this change prevents the formation of the stable triclosan-NAD+-FabI complex, and both this alteration and its overexpression contributed to achieving MICs of 1 to 2 μg/ml for these strains. Three novel, potent inhibitors of FabI (50% inhibitory concentrations, ≤64 nM) demonstrated up to 1,000-fold better activity than triclosan against the strains for which triclosan MICs were elevated. None of the compounds tested from this series formed a stable complex with NAD+-FabI. Consequently, although the overexpression of wild-type FabI gave rise to an increase in the MICs, as expected, overexpression of FabI with an F204C alteration did not cause an additional increase in resistance. Therefore, this work identifies the mechanisms of triclosan resistance in S. aureus, and we present three compounds from a novel chemical series of FabI inhibitors which have excellent activities against both triclosan-resistant and -sensitive clinical isolates of S. aureus.

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Antibacterial Activity of N-Pentylpantothenamide Is Due to Inhibition of Coenzyme A Synthesis

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.01473-09 ◽

2010 ◽

Vol 54 (3) ◽

pp. 1374-1377 ◽

Cited By ~ 27

Author(s):

Jacob Thomas ◽

John E. Cronan

Keyword(s):

Antibacterial Activity ◽

Growth Inhibition ◽

Coenzyme A ◽

Acyl Carrier Protein ◽

Carrier Protein ◽

Prosthetic Group ◽

Protein Moiety

ABSTRACT Growth inhibition by the pantothenate analog N-pentylpantothenamide (N5-Pan) has been attributed to the accumulation of acyl carrier protein carrying a prosthetic group modified by incorporation of N5-Pan. This was attributed to an inability of the AcpH acyl carrier protein phosphodiesterase to cleave the N5-Pan-modified prosthetic group from the protein moiety. We report that AcpH readily removes the N5-Pan-modified prosthetic group both in vivo and in vitro and show that N5-Pan blocks coenzyme A synthesis.

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Purification, Characterization, and Identification of Novel Inhibitors of the β-Ketoacyl-Acyl Carrier Protein Synthase III (FabH) from Staphylococcus aureus

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.46.5.1310-1318.2002 ◽

2002 ◽

Vol 46 (5) ◽

pp. 1310-1318 ◽

Cited By ~ 36

Author(s):

Xin He ◽

Kevin A. Reynolds

Keyword(s):

Staphylococcus Aureus ◽

Fatty Acid ◽

Fatty Acid Synthase ◽

Fatty Acid Biosynthesis ◽

Genomic Sequence ◽

Acyl Carrier Protein ◽

Polyacrylamide Gel Electrophoresis ◽

Sodium Dodecyl ◽

Size Exclusion ◽

Carrier Protein

ABSTRACT Staphylococcus aureus is a versatile and dangerous pathogen and one of the major causes of community-acquired and hospital-acquired infections. The rise of multidrug-resistant strains of S. aureus requires the development of new antibiotics with previously unexploited mechanisms of action, such as inhibition of the β-ketoacyl-acyl carrier protein (ACP) synthase III (FabH). This enzyme initiates fatty acid biosynthesis in a bacterial type II fatty acid synthase, catalyzing a decarboxylative condensation between malonyl-ACP and an acyl coenzyme A (CoA) substrate and is essential for viability. We have identified only one fabH in the genome of S. aureus and have shown that it encodes a protein with 57, 40, and 34% amino acid sequence identity with the FabH proteins of Bacillus subtilis (bFabH1), Escherichia coli (ecFabH), and Mycobacterium tuberculosis (mtFabH). Additional genomic sequence analysis revealed that this S. aureus FabH (saFabH) is not mutated in certain methicillin-resistant S. aureus (MRSA) and vancomycin-resistant S. aureus (VRSA) strains. saFabH was expressed in E. coli with an N-terminal polyhistidine tag and subsequently purified by metal chelate and size exclusion chromatography. Analysis by sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed a molecular mass of 37 kDa, while gel filtration demonstrated a mass of 66.7 kDa, suggesting a noncovalent homodimeric structure for saFabH. The apparent Km for malonyl-ACP was 1.76 ± 0.40 μM, and the enzyme was active with acetyl-CoA (k cat, 16.18 min−1; Km , 6.18 ± 0.9 μM), butyryl-CoA (k cat, 42.90 min−1; Km , 2.32 ± 0.12 μM), and isobutyryl-CoA (k cat, 98.0 min−1; Km , 0.32 ± 0.04 μM). saFabH was weakly inhibited by thiolactomycin (50% inhibitory concentration [IC50], >100 μM) yet was efficiently inhibited by two new FabH inhibitors, 5-chloro-4-phenyl-[1,2]-dithiol-3-one (IC50, 1.87 ± 0.10 μM) and 4-phenyl-5-phenylimino-[1,2,4]dithiazolidin-3-one (IC50, 0.775 ± 0.08 μM).

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