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

He, Xin; Reynolds, Kevin A.

doi:10.1128/aac.46.5.1310-1318.2002

Cited by 55 publications

(69 citation statements)

References 41 publications

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“…2B). These residues include the catalytic nucleophile Cys112 (E. coli FabH numbering); the general base His244, which also binds the carbonyl oxygen of the malonyl-ACP extender unit; Asn274, which promotes the decarboxylation of malonyl-CoA via stabilization of the enol intermediate; Gly306, which stabilizes the oxyanion; and Arg42, which takes part in maintaining the structural integrity of the ligand binding domain (28). Residues that are implicated in interactions either with the pantheteine arm of the holo-ACP (i.e., Asn247, Gly209, Ile156, Met207, Val212, Phe213, and Ile250) or with the adenine ring of the coenzyme A (CoA) (Trp32 and Arg151) are not conserved.…”

Section: Resultsmentioning

confidence: 99%

“…In contrast, FabH from Staphylococcus aureus, which produces a relatively high percentage of branched-chain fatty acids, accepts acetyl-CoA and butyryl-CoA as primers, but the most preferred substrate is isobutyryl-CoA. Although the substrate specificities of the S. aureus and E. coli FabH proteins are dramatically different, amino acids within the substrate binding pockets of these enzymes are highly conserved (28).…”

Section: Vol 185 2003 Dar Biosynthesis 865mentioning

confidence: 95%

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2,5-Dialkylresorcinol Biosynthesis in Pseudomonas aurantiaca : Novel Head-to-Head Condensation of Two Fatty Acid-Derived Precursors

et al. 2003

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2-Hexyl-5-propylresorcinol is the predominant analog of several dialkylresorcinols produced by Pseudomonas aurantiaca (Pseudomonas fluorescens BL915). We isolated and characterized three biosynthetic genes that encode an acyl carrier protein, a ␤-ketoacyl-acyl carrier protein synthase III, and a protein of unknown function, all of which collectively allow heterologous production of 2-hexyl-5-propylresorcinol in Escherichia coli. Two regulatory genes exhibiting similarity to members of the AraC family of transcriptional regulators are also present in the identified gene cluster. Based on the deduced functions of the proteins encoded by the gene cluster and the observed incorporation of labeled carbons from octanoic acid into 2-hexyl-5-propylresorcinol, we propose that dialkylresorcinols are derived from medium-chain-length fatty acids by an unusual head-tohead condensation of ␤-ketoacyl thioester intermediates. Genomic evidence suggests that there is a similar pathway for the biosynthesis of the flexirubin-type pigments in certain bacteria belonging to the order Cytophagales.

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Section: Resultsmentioning

confidence: 99%

Section: Vol 185 2003 Dar Biosynthesis 865mentioning

confidence: 95%

2,5-Dialkylresorcinol Biosynthesis in Pseudomonas aurantiaca : Novel Head-to-Head Condensation of Two Fatty Acid-Derived Precursors

et al. 2003

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“…Consistent with this observation, the Streptomyces glaucescens FabH (sgFabH) was shown to have less stringent substrate specificity than the ecFabH (6), utilizing both straight-and branched-chain acyl-CoAs. Subsequent analyses revealed that FabH enzymes from other organisms which produce both BCFAs and SCFAs had similar substrate flexibilities (1,8).…”

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confidence: 99%

Alteration of the Fatty Acid Profile of Streptomyces coelicolor by Replacement of the Initiation Enzyme 3-Ketoacyl Acyl Carrier Protein Synthase III (FabH)

Florova

Reynolds

2005

J Bacteriol

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The first elongation step of fatty acid biosynthesis by a type II dissociated fatty acid synthases is catalyzed by 3-ketoacyl-acyl carrier protein (ACP) synthase III (KASIII, FabH). This enzyme, encoded by the fabH gene, catalyzes a decarboxylative condensation between an acyl coenzyme A (CoA) primer and malonyl-ACP. In organisms such as Escherichia coli, which generate only straight-chain fatty acids (SCFAs), FabH has a substrate preference for acetyl-CoA. In streptomycetes and other organisms which produce a mixture of both SCFAs and branched-chain fatty acids (BCFAs), FabH has been shown to utilize straight-and branched-chain acyl-CoA substrates. We report herein the generation of a Streptomyces coelicolor mutant (YL/ecFabH) in which the chromosomal copy of the fabH gene has been replaced and the essential process of fatty acid biosynthesis is initiated by plasmid-based expression of the E. coli FabH (bearing only 35% amino acid identity to the Streptomyces enzyme). The YL/ecFabH mutant produces predominantly SCFAs (86%). In contrast, BCFAs predominate (ϳ70%) in both the S. coelicolor parental strain and S. coelicolor YL/sgFabH (a ⌬fabH mutant carrying a plasmid expressing the Streptomyces glaucescens FabH). These results provide the first unequivocal evidence that the substrate specificity of FabH observed in vitro is a determinant of the fatty acid made in an organism. The YL/ecFabH strain grows significantly slower on both solid and liquid media. The levels of FabH activity in cell extracts of YL/ecFabH were also significantly lower than those in cell extracts of YL/sgFabH, suggesting that a decreased rate of fatty acid synthesis may account for the observed decreased growth rate. The production of low levels of BCFAs in YL/ecFabH suggests either that the E. coli FabH is more tolerant of different acyl-CoAs substrates than previously thought or that there is an additional pathway for initiation of BCFA biosynthesis in Streptomyces coelicolor.

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“…We describe herein a separate approach in which we screened the National Cancer Institute (NCI) database for compounds that possess some of the structural characteristics of TLM which have been shown to be important for inhibition of the condensing enzymes of type II FASs. This work has led to the identification of substituted 1,2-dithiole-3-ones, which preliminary analyses have demonstrated to be potent inhibitors of ecFabH, saFabH, and pfFabH (27,53). A series of analogs based on this basic skeleton have been prepared and tested against both ecFabH and saFabH.…”

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confidence: 99%

“…Initial work focused on Escherichia coli FabH (ecFabH) (64) but has been extended to include FabH proteins from important human pathogens, such as Mycobacterium tuberculosis (mtFabH) (14,59), Staphylococcus aureus (saFabH) (27), Streptococcus pneumoniae (spFabH) (34), and Plasmodium falciparum (pfFabH) (53). In all cases, FabH catalyzes a direct condensation between the acyl coenzyme A (CoA) primer used to initiate fatty acid biosynthesis and MACP (13,14,16,25,64).…”

mentioning

confidence: 99%

1,2-Dithiole-3-Ones as Potent Inhibitors of the Bacterial 3-Ketoacyl Acyl Carrier Protein Synthase III (FabH)

Reeve

Desai

et al. 2004

Antimicrob Agents Chemother

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The enzyme FabH catalyzes the initial step of fatty acid biosynthesis via a type II dissociated fatty acid synthase. The pivotal role of this essential enzyme, combined with its unique structural features and ubiquitous occurrence in bacteria, has made it an attractive new target for the development of antibacterial and antiparasitic compounds. We have searched the National Cancer Institute database for compounds bearing structural similarities to thiolactomycin, a natural product which exhibits a weak activity against FabH. This search has yielded several substituted 1,2-dithiole-3-ones that are potent inhibitors of FabH from both Escherichia coli (ecFabH) and Staphylococcus aureus (saFabH). The most potent inhibitor was 4,5-dichloro-1,2-dithiole-3-one, which had 50% inhibitory concentration (IC 50 ) values of 2 M (ecFabH) and 0.16 M (saFabH). The corresponding 3-thione analog exhibited comparable activities. Analogs in which the 4-chloro substituent was replaced with a phenyl group were also potent inhibitors, albeit somewhat less effectively (IC 50 values of 5.7 and 0.98 M for ecFabH and saFabH, respectively). All of the 5-chlorinated inhibitors were most effective when they were preincubated with FabH in the absence of substrates. The resulting enzyme-inhibitor complex did not readily regain activity after excess inhibitor was removed, suggesting that a slow dissociation occurs. In stark contrast, a series of inhibitors in which the 5-chloro substituent was replaced with the isosteric and isoelectronic trifluoromethyl group were poorer inhibitors (IC 50 values typically ranging from 25 to >100 M for both ecFabH and saFabH), did not require a preincubation period for maximal activity, and generated an enzymeinhibitor complex which readily dissociated. Possible modes of binding of 5-chloro-1,2-dithiole-3-ones and 5-chloro-1,2-dithiole-3-thiones with FabH which account for the role of the 5-chloro substituent were considered.Fatty acid biosynthesis, an essential process for all organisms, is catalyzed in plants and bacteria by a series of discrete dissociable enzymes and a central acyl carrier protein (ACP) (43). This set of enzymes is known collectively as a type II fatty acid synthase (FAS) and differs significantly from the type I FAS of metazoans, in which all of the enzymatic activities are contained on one or two polypeptides (12,31,67). The structural and mechanistic differences between the two FAS systems, in conjunction with the fact that the type I FAS is down regulated in well-nourished mammals (38, 39), have led to significant interest in components of the type II FAS as targets for the development of new antibacterial agents. Such agents may also have promise as novel antimalarials because protozoan parasites of the genus Plasmodium have been shown recently to contain a type II FAS in their apicoplast (53,62,63).Significant efforts and progress have been made in the understanding of small-molecule inhibition of two different components of the type II FAS. FabI (InhA), the enoyl ACP reductase that c...

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

Cited by 55 publications

References 41 publications

2,5-Dialkylresorcinol Biosynthesis in Pseudomonas aurantiaca : Novel Head-to-Head Condensation of Two Fatty Acid-Derived Precursors

2,5-Dialkylresorcinol Biosynthesis in Pseudomonas aurantiaca : Novel Head-to-Head Condensation of Two Fatty Acid-Derived Precursors

Alteration of the Fatty Acid Profile of Streptomyces coelicolor by Replacement of the Initiation Enzyme 3-Ketoacyl Acyl Carrier Protein Synthase III (FabH)

1,2-Dithiole-3-Ones as Potent Inhibitors of the Bacterial 3-Ketoacyl Acyl Carrier Protein Synthase III (FabH)

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