Products of fatty acid synthesis by a particulate fraction from germinating pea (Pisum sativum L.)

The synthesis of lipids and acyl thioesters was studied in microsomal preparations from germinating pea (Pisum sativum cv. Feltham First) seeds. Under conditions of maximal synthesis (in the presence of exogenous acyl-carrier protein) acyl-acyl-carrier proteins accounted for about half the total incorporation from [14C]malonyl-CoA. Decreasing the concentrations of exogenous acyl-carrier protein lowered the overall synthesis of fatty acids by decreasing, almost exclusively, the radioactivity associated with acyl-acyl-carrier proteins. A time-course experiment showed that acyl-acyl-carrier proteins accumulated most of the radioactive label at the beginning of the incubation but, eventually, the amount of radioactivity in that fraction decreased, while a simultaneous increase in the acyl-CoA and lipid fractions was noticed. Addition of exogenous CoA (1 mM) produced a decrease of total incorporation, but an increase in the radioactivity incorporated into acyl-CoA. The microsomal preparations synthesized saturated fatty acids up to C20, including significant proportions of pentadecanoic acid and heptadecanoic acid. Synthesis of these ‘odd-chain’ fatty acids only took place in the microsomal fraction. In contrast, when the 18,000g supernatant (containing the microsomal and soluble fractions) was incubated with [14C]malonyl-CoA, the radioactive fatty acid and acyl classes closely resembled the patterns produced by germinating in the presence of [14C]acetate in vivo. The results are discussed in relation to the role of acyl thioesters in the biosynthesis of plant lipids.

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Concentrations of long-chain acyl-acyl carrier proteins during fatty acid synthesis by chloroplasts isolated from pea (Pisum sativum), safflower (Carthamus tinctoris) and amaranthus (Amaranthus lividus) leaves

Archives of Biochemistry and Biophysics ◽

10.1016/0003-9861(90)90007-l ◽

1990 ◽

Vol 276 (1) ◽

pp. 38-46 ◽

Cited By ~ 8

Author(s):

Grattan Roughan ◽

Ikuo Nishida

Keyword(s):

Fatty Acid ◽

Pisum Sativum ◽

Fatty Acid Synthesis ◽

Acid Synthesis ◽

Carrier Proteins ◽

Acyl Carrier Proteins ◽

Chain Acyl ◽

Long Chain

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The Two Functional Enoyl-Acyl Carrier Protein Reductases of Enterococcus faecalis Do Not Mediate Triclosan Resistance

mBio ◽

10.1128/mbio.00613-13 ◽

2013 ◽

Vol 4 (5) ◽

Cited By ~ 26

Author(s):

Lei Zhu ◽

Hongkai Bi ◽

Jincheng Ma ◽

Zhe Hu ◽

Wenbin Zhang ◽

...

Keyword(s):

Fatty Acids ◽

Fatty Acid ◽

Enterococcus Faecalis ◽

Fatty Acid Synthesis ◽

Acyl Carrier Protein ◽

Acid Synthesis ◽

Carrier Protein ◽

Content Type ◽

Enoyl Acp Reductase ◽

Triclosan Resistance

ABSTRACTEnoyl-acyl carrier protein (enoyl-ACP) reductase catalyzes the last step of the elongation cycle in the synthesis of bacterial fatty acids. TheEnterococcus faecalisgenome contains two genes annotated as enoyl-ACP reductases, a FabI-type enoyl-ACP reductase and a FabK-type enoyl-ACP reductase. We report that expression of either of the two proteins restores growth of anEscherichia colifabItemperature-sensitive mutant strain under nonpermissive conditions.In vitroassays demonstrated that both proteins support fatty acid synthesis and are active with substrates of all fatty acid chain lengths. Although expression ofE. faecalis fabKconfers toE. colihigh levels of resistance to the antimicrobial triclosan, deletion offabKfrom theE. faecalisgenome showed that FabK does not play a detectable role in the inherent triclosan resistance ofE. faecalis. Indeed, FabK seems to play only a minor role in modulating fatty acid composition. Strains carrying a deletion offabKgrow normally without fatty acid supplementation, whereasfabIdeletion mutants make only traces of fatty acids and are unsaturated fatty acid auxotrophs.IMPORTANCEThe finding that exogenous fatty acids support growth ofE. faecalisstrains defective in fatty acid synthesis indicates that inhibitors of fatty acid synthesis are ineffective in counteringE. faecalisinfections because host serum fatty acids support growth of the bacterium.

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Overproduction of a Functional Fatty Acid Biosynthetic Enzyme Blocks Fatty Acid Synthesis inEscherichia coli

Journal of Bacteriology ◽

10.1128/jb.180.17.4596-4602.1998 ◽

1998 ◽

Vol 180 (17) ◽

pp. 4596-4602 ◽

Cited By ~ 54

Author(s):

Satyanarayana Subrahmanyam ◽

John E. Cronan

Keyword(s):

Fatty Acid ◽

Fatty Acid Synthesis ◽

Acyl Carrier Protein ◽

State Level ◽

Acid Synthesis ◽

The Other ◽

Carrier Protein ◽

Biosynthetic Enzyme ◽

E Coli ◽

Malonyl Coa

ABSTRACT β-Ketoacyl-acyl carrier protein (ACP) synthetase II (KAS II) is one of three Escherichia coli isozymes that catalyze the elongation of growing fatty acid chains by condensation of acyl-ACP with malonyl-ACP. Overexpression of this enzyme has been found to be extremely toxic to E. coli, much more so than overproduction of either of the other KAS isozymes, KAS I or KAS III. The immediate effect of KAS II overproduction is the cessation of phospholipid synthesis, and this inhibition is specifically due to the blockage of fatty acid synthesis. To determine the cause of this inhibition, we examined the intracellular pools of ACP, coenzyme A (CoA), and their acyl thioesters. Although no significant changes were detected in the acyl-ACP pools, the CoA pools were dramatically altered by KAS II overproduction. Malonyl-CoA increased to about 40% of the total cellular CoA pool upon KAS II overproduction from a steady-state level of around 0.5% in the absence of KAS II overproduction. This finding indicated that the conversion of malonyl-CoA to fatty acids had been blocked and could be explained if either the conversion of malonyl-CoA to malonyl-ACP and/or the elongation reactions of fatty acid synthesis had been blocked. Overproduction of malonyl-CoA:ACP transacylase, the enzyme catalyzing the conversion of malonyl-CoA to malonyl-ACP, partially relieved the toxicity of KAS II overproduction, consistent with a model in which high levels of KAS II blocks access of the other KAS isozymes to malonyl-CoA:ACP transacylase.

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Enoyl-Acyl Carrier Protein Reductase I (FabI) Is Essential for the Intracellular Growth of Listeria monocytogenes

Infection and Immunity ◽

10.1128/iai.00647-16 ◽

2016 ◽

Vol 84 (12) ◽

pp. 3597-3607 ◽

Cited By ~ 8

Author(s):

Jiangwei Yao ◽

Megan E. Ericson ◽

Matthew W. Frank ◽

Charles O. Rock

Keyword(s):

Fatty Acids ◽

Fatty Acid ◽

Fatty Acid Synthesis ◽

Acyl Carrier Protein ◽

Acid Synthesis ◽

Carrier Protein ◽

Type Ii ◽

Intracellular Growth ◽

Content Type ◽

Bacterial Fatty Acid

Enoyl-acyl carrier protein reductase catalyzes the last step in each elongation cycle of type II bacterial fatty acid synthesis and is a key regulatory protein in bacterial fatty acid synthesis. Genes of the facultative intracellular pathogenListeria monocytogenesencode two functional enoyl-acyl carrier protein isoforms based on their ability to complement the temperature-sensitive growth phenotype ofEscherichia colistrain JP1111 [fabI(Ts)]. The FabI isoform was inactivated by the FabI selective inhibitor AFN-1252, but the FabK isoform was not affected by the drug, as expected. Inhibition of FabI by AFN-1252 decreased endogenous fatty acid synthesis by 80% and lowered the growth rate ofL. monocytogenesin laboratory medium. Robust exogenous fatty acid incorporation was not detected inL. monocytogenesunless the pathway was partially inactivated by AFN-1252 treatment. However, supplementation with exogenous fatty acids did not restore normal growth in the presence of AFN-1252. FabI inactivation prevented the intracellular growth ofL. monocytogenes, showing that neither FabK nor the incorporation of host cellular fatty acids was sufficient to support the intracellular growth ofL. monocytogenes. Our results show that FabI is the primary enoyl-acyl carrier protein reductase of type II bacterial fatty acid synthesis and is essential for the intracellular growth ofL. monocytogenes.

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Fatty acid biosynthesis by a particulate preparation from germinating pea

Biochemical Journal ◽

10.1042/bj1680261 ◽

1977 ◽

Vol 168 (2) ◽

pp. 261-269 ◽

Cited By ~ 45

Author(s):

Paul Bolton ◽

John L. Harwood

Keyword(s):

Fatty Acids ◽

Fatty Acid ◽

Stearic Acid ◽

Palmitic Acid ◽

Acyl Carrier Protein ◽

Acid Synthesis ◽

Long Chain Fatty Acids ◽

Dipalmitoyl Phosphatidylcholine ◽

Malonyl Coa ◽

Long Chain

1. Fatty acid synthesis was studied in microsomal preparations from germinating pea (Pisum sativum). 2. The preparations synthesized a mixture of saturated fatty acids up to a chain length of C24 from [14C]malonyl-CoA. 3. Whereas hexadecanoic acid was made de novo, octadecanoic acid and icosanoic acid were synthesized by elongation. 4. The products formed during [14C]malonyl-CoA incubation were analysed, and unesterified fatty acids and polar lipids were found to be major products. [14C]Palmitic acid represented a high percentage of the acyl-carrier protein esters, whereas 14C-labelled very-long-chain fatty acids were mainly present as unesterified fatty acids. CoA esters were minor products. 5. The addition of exogenous lipids to the incubation system usually resulted in stimulation of [14C]malonyl-CoA incorporation into fatty acids. The greatest stimulation was obtained with dipalmitoyl phosphatidylcholine. Both exogenous palmitic acid and dipalmitoyl phosphatidylcholine increased the amount of [14C]-stearic acid synthesized, relative to [14C]palmitic acid. Addition of stearic acid increased the amount of [14C]icosanoic acid formed. 6. [14C]Stearic acid was elongated more effectively to icosanoic acid than [14C]stearoyl-CoA, and its conversion was not decreased by addition of unlabelled stearoyl-CoA. 7. Incorporation of [14C]malonyl-CoA into fatty acids was markedly decreased by iodoacetamide and 5,5′-dithiobis-(2-nitrobenzoic acid). Palmitate elongation was sensitive to arsenite addition, and stearate elongation to the presence of Triton X-100 or fluoride. The action of fluoride was not, apparently, due to chelation. 8. The microsomal preparations differed from soluble fractions from germinating pea in (a) synthesizing very-long-chain fatty acids, (b) not utilizing exogenous palmitate–acyl-carrier protein as a substrate for palmitate elongation and (c) having fatty acid synthesis stimulated by the addition of certain complex lipids.

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The fractionation of the fatty acid synthetase activities of avocado mesocarp plastids

Biochemical Journal ◽

10.1042/bj1460439 ◽

1975 ◽

Vol 146 (2) ◽

pp. 439-445 ◽

Cited By ~ 29

Author(s):

P J Weaire ◽

R G O Kekwick

Keyword(s):

Fatty Acids ◽

Fatty Acid ◽

De Novo ◽

Acyl Carrier Protein ◽

Fatty Acid Synthetase ◽

Carrier Protein ◽

Acetyl Coa ◽

Principal Product ◽

Absolute Requirement ◽

Malonyl Coa

1. The range of fatty acids formed by preparations of ultrasonically ruptured avocado mesocarp plastids was dependent on the substrate. Whereas [1-14C]palmitate and [14C]oleate were the major products obtained from [-14C]acetate and [1-14C]acetyl-CoA, the principal product from [2-14C]malonyl-CoA was [14-C]stearate. 2. Ultracentrifugation of the ruptured plastids at 105000g gave a supernatant that formed mainly stearate from [2-14C]malonyl-CoA and to a lesser extent from [1-14C]acetate. The incorporation of [1-14C]acetate into stearate by this fraction was inhibited by avidin. 3. The 105000g precipitate of the disrupted plastids incorporated [1-14C]acetate into a mixture of fatty acids that contained largely [14C]plamitate and [14C]oleate. The formation of [14C]palmitate and [14C]oleate by disrupted plastids was unaffected by avidin. 4. The soluble fatty acid synthetase was precipitated from the 105000g supernatant in the 35-65%-saturated-(NH4)2SO4 fraction and showed an absolute requirement for acyl-carrier protein. 5. Both fractions synthesized fatty acids de novo.

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