FadD from Pseudomonas putida CA-3 Is a True Long-Chain Fatty Acyl Coenzyme A Synthetase That Activates Phenylalkanoic and Alkanoic Acids

ABSTRACT A fatty acyl coenzyme A synthetase (FadD) from Pseudomonas putida CA-3 is capable of activating a wide range of phenylalkanoic and alkanoic acids. It exhibits the highest rates of reaction and catalytic efficiency with long-chain aromatic and aliphatic substrates. FadD exhibits higher k cat and Km values for aromatic substrates than for the aliphatic equivalents (e.g., 15-phenylpentadecanoic acid versus pentadecanoic acid). FadD is inhibited noncompetitively by both acrylic acid and 2-bromooctanoic acid. The deletion of the fadD gene from P. putida CA-3 resulted in no detectable growth or polyhydroxyalkanoate (PHA) accumulation with 10-phenyldecanoic acid, decanoic acid, and longer-chain substrates. The results suggest that FadD is solely responsible for the activation of long-chain phenylalkanoic and alkanoic acids. While the CA-3ΔfadD mutant could grow on medium-chain substrates, a decrease in growth yield and PHA accumulation was observed. The PHA accumulated by CA-3ΔfadD contained a greater proportion of short-chain monomers than did wild-type PHA. Growth of CA-3ΔfadD was unaffected, but PHA accumulation decreased modestly with shorter-chain substrates. The complemented mutant regained 70% to 90% of the growth and PHA-accumulating ability of the wild-type strain depending on the substrate. The expression of an extra copy of fadD in P. putida CA-3 resulted in increased levels of PHA accumulation (up to 1.6-fold) and an increase in the incorporation of longer-monomer units into the PHA polymer.

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Long-Chain Fatty Acyl Coenzyme A Ligase FadD2 Mediates Intrinsic Pyrazinamide Resistance in Mycobacterium tuberculosis

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.02130-16 ◽

2016 ◽

Vol 61 (2) ◽

Cited By ~ 8

Author(s):

Brandon C. Rosen ◽

Nicholas A. Dillon ◽

Nicholas D. Peterson ◽

Yusuke Minato ◽

Anthony D. Baughn

Keyword(s):

Mycobacterium Tuberculosis ◽

Coenzyme A ◽

Fatty Acyl ◽

Wild Type ◽

Intrinsic Resistance ◽

First Line ◽

Minimum Concentration ◽

Content Type ◽

Acyl Coenzyme A ◽

Long Chain

ABSTRACT Pyrazinamide (PZA) is a first-line tuberculosis (TB) drug that has been in clinical use for 60 years yet still has an unresolved mechanism of action. Based upon the observation that the minimum concentration of PZA required to inhibit the growth of Mycobacterium tuberculosis is approximately 1,000-fold higher than that of other first-line drugs, we hypothesized that M. tuberculosis expresses factors that mediate intrinsic resistance to PZA. To identify genes associated with intrinsic PZA resistance, a library of transposon-mutagenized Mycobacterium bovis BCG strains was screened for strains showing hypersusceptibility to the active form of PZA, pyrazinoic acid (POA). Disruption of the long-chain fatty acyl coenzyme A (CoA) ligase FadD2 enhanced POA susceptibility by 16-fold on agar medium, and the wild-type level of susceptibility was restored upon expression of fadD2 from an integrating mycobacterial vector. Consistent with the recent observation that POA perturbs mycobacterial CoA metabolism, the fadD2 mutant strain was more vulnerable to POA-mediated CoA depletion than the wild-type strain. Ectopic expression of the M. tuberculosis pyrazinamidase PncA, necessary for conversion of PZA to POA, in the fadD2 transposon insertion mutant conferred at least a 16-fold increase in PZA susceptibility under active growth conditions in liquid culture at neutral pH. Importantly, deletion of fadD2 in M. tuberculosis strain H37Rv also resulted in enhanced susceptibility to POA. These results indicate that FadD2 is associated with intrinsic PZA and POA resistance and provide a proof of concept for the target-based potentiation of PZA activity in M. tuberculosis.

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Long-chain fatty acyl-coenzyme A suppresses hepatitis C virus infection by targeting virion-bound lipoproteins

Antiviral Research ◽

10.1016/j.antiviral.2020.104734 ◽

2020 ◽

Vol 177 ◽

pp. 104734 ◽

Cited By ~ 2

Author(s):

Xinlei Li ◽

Jinqian Li ◽

Yetong Feng ◽

Hua Cai ◽

Yi-Ping Li ◽

...

Keyword(s):

Hepatitis C Virus ◽

Hepatitis C ◽

Virus Infection ◽

Coenzyme A ◽

Fatty Acyl ◽

Hepatitis C Virus Infection ◽

Acyl Coenzyme A ◽

Long Chain

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Steady-state concentrations of coenzyme A, acetyl-coenzyme A and long-chain fatty acyl-coenzyme A in rat-liver mitochondria oxidizing palmitate

Biochemical Journal ◽

10.1042/bj0970587 ◽

1965 ◽

Vol 97 (2) ◽

pp. 587-594 ◽

Cited By ~ 206

Author(s):

PB Garland ◽

D Shepherd ◽

DW Yates

Keyword(s):

Rat Liver ◽

Coenzyme A ◽

Liver Mitochondria ◽

Fatty Acyl ◽

Rat Liver Mitochondria ◽

Beta Oxidation ◽

Acetyl Coa ◽

Acetyl Coenzyme A ◽

Acyl Coenzyme A ◽

Long Chain

1. Fluorimetric assays are described for CoASH, acetyl-CoA and long-chain fatty acyl-CoA, and are sensitive to at least 50mumumoles of each. 2. Application of these assays to rat-liver mitochondria oxidizing palmitate in the absence and presence of carnitine indicated two pools of intramitochondrial CoA. One pool could be acylated by palmitate and ATP, and the other pool acylated by palmitate with ATP and carnitine, or by palmitoylcarnitine alone. 3. The intramitochondrial content of acetyl-CoA is increased by the oxidation of palmitate both in the absence and presence of l-malate. 4. The conversion of palmitoyl-CoA into acetyl-CoA by beta-oxidation takes place without detectable accumulation of acyl-CoA intermediates.

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