scholarly journals Fatty acid biosynthesis by a particulate preparation from germinating pea

1977 ◽  
Vol 168 (2) ◽  
pp. 261-269 ◽  
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.

Chloroacetamide Mode of Action, I: Inhibition of Very Long Chain Fatty Acid Synthesis in Scenedesmus acutus

1998 ◽  
Vol 53 (11-12) ◽  
pp. 995-1003 ◽  
Keyword(s):  
Fatty Acids ◽  
Cell Wall ◽  
Oleic Acid ◽  
Fatty Acid ◽  
Acid Synthesis ◽  
Resistant Cell Line ◽  
Long Chain Fatty Acids ◽  
Scenedesmus Acutus ◽  
Long Chain ◽  
Chain Fatty Acids

Abstract Herbicidal chloroacetamides cause a very sensitive inhibition of fatty acid incorporation into an insoluble cell wall fraction of Scenedesmus acutus. The molecular basis was investigated in more detail. After incubation of the algae with [14C]oleic acid and saponification, the remaining pellet was solubilized and fractionated consecutively with chloroform / methanol, phosphate buffer, amylase, pronase, and finally with dioxane/HCl. By acid hydrolysis in dioxane a part of the cell wall residue was solubilized showing inhibition of exogenously applied oleic acid and other labelled precursors such as stearic acid, palmitic acid, and acetate. After extraction of this dioxane-soluble subfraction with hexane, HPLC could separate labelled metabolites less polar than oleic acid. T heir formation was completely inhibited by chloroacetam ides, e.g. 1 μᴍ metazachlor. This effect was also observed with the herbicidally active 5-enantiomer of metolachlor while the inactive R-enantiomer had no influence. These strongly inhibited metabolites could be characterized by radio-HPLC /MS as very long chain fatty acids (VLCFAs) with a carbon chain between 20 and 26. Incubating am etazachlor-resistant cell line of S. acutus (Mz-1) with [14C]oleic acid, V LCFA s could not be detected in the dioxane/ HCl-subfraction. Furthermore, comparing the presence of endogenous fatty acids in wildtype and mutant Mz-1 the VLCFA content of the mutant is very low, while the content of long chain fatty acids (C16 -18) is increased, particularly oleic acid. Obviously, the phytotoxicity of chloroacetam ides in S. acutus is due to inhibition of VLCFA synthesis. The resistance of the mutant to metazachlor has a bearing on the higher amount of long chain fatty acids replacing the missing VLCFAs in essential membranes or cell wall components.

The Very-Long-Chain Fatty Acid Synthase Is Inhibited by Chloroacetamides

2004 ◽  
Vol 59 (7-8) ◽  
pp. 549-553 ◽  
Author(s):  
Thomas Götz ◽  
Peter Böger
Keyword(s):  
Fatty Acids ◽  
Fatty Acid ◽  
Fatty Acid Synthase ◽  
Chain Fatty Acid ◽  
Long Chain Fatty Acids ◽  
Irreversible Binding ◽  
Long Chain Fatty Acid ◽  
Malonyl Coa ◽  
Elongation Step ◽  
Long Chain

AbstractThe first elongation step to form very-long-chain fatty acids (VLCFAs) is catalyzed by the VLCFA-synthase. CoA-activated fatty acids react with malonyl-CoA to condense a C2-unit. As shown with recombinant enzyme this reaction is specifically inhibited by chloroacetamide herbicides. The inhibition is alleviated when the inhibitor (e.g. metazachlor) is incubated together with adequate concentrations of the substrate (e.g. oleoyl-CoA). Malonyl-CoA has no influence. However, once a chloroacetamide has been tightly bound to the synthase after an appropriate time it cannot be displaced anymore by the substrate. In contrast, oleoyl- CoA, is easily removed from the synthase by metazachlor. The irreversible binding of the chloroacetamides and their competition with the substrate explains the very low half-inhibition values of 10-8 м and below. Chiral chloroacetamides like metolachlor or dimethenamid give identical results. However, only the (S)-enantiomers are active.

scholarly journals Identification and biophysical characterization of a very-long-chain-fatty-acid-substituted phosphatidylinositol in yeast subcellular membranes

2004 ◽  
Vol 381 (3) ◽  
pp. 941-949 ◽  
Author(s):  
Roger SCHNEITER ◽  
Britta BRÜGGER ◽  
Clare M. AMANN ◽  
Glenn D. PRESTWICH ◽  
Raquel F. EPAND ◽  
...  
Keyword(s):  
Fatty Acids ◽  
Fatty Acid ◽  
Lipid Analysis ◽  
Hexagonal Phase ◽  
Acid Synthesis ◽  
Long Chain Fatty Acids ◽  
Biophysical Properties ◽  
Ceramide Synthesis ◽  
Long Chain ◽  
Chain Fatty Acids

Morphological analysis of a conditional yeast mutant in acetyl-CoA carboxylase acc1ts/mtr7, the rate-limiting enzyme of fatty acid synthesis, suggested that the synthesis of C26 VLCFAs (very-long-chain fatty acids) is important for maintaining the structure and function of the nuclear membrane. To characterize this C26-dependent pathway in more detail, we have now examined cells that are blocked in pathways that require C26. In yeast, ceramide synthesis and remodelling of GPI (glycosylphosphatidylinositol)-anchors are two pathways that incorporate C26 into lipids. Conditional mutants blocked in either ceramide synthesis or the synthesis of GPI anchors do not display the characteristic alterations of the nuclear envelope observed in acc1ts, indicating that the synthesis of another C26-containing lipid may be affected in acc1ts mutant cells. Lipid analysis of isolated nuclear membranes revealed the presence of a novel C26-substituted PI (phosphatidylinositol). This C26-PI accounts for approx. 1% of all the PI species, and is present in both the nuclear and the plasma membrane. Remarkably, this C26-PI is the only C26-containing glycerophospholipid that is detectable in wild-type yeast, and the C26-substitution is highly specific for the sn-1 position of the glycerol backbone. To characterize the biophysical properties of this lipid, it was chemically synthesized. In contrast to PIs with normal long-chain fatty acids (C16 or C18), the C26-PI greatly reduced the bilayer to hexagonal phase transition of liposomes composed of 1,2-dielaidoyl-sn-glycero-3-phosphoethanolamine (DEPE). The biophysical properties of this lipid are thus consistent with a possible role in stabilizing highly curved membrane domains.

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

1981 ◽  
Vol 199 (1) ◽  
pp. 221-226 ◽  
Author(s):  
J Sanchez ◽  
J L Harwood
Keyword(s):  
Fatty Acids ◽  
Fatty Acid ◽  
Pisum Sativum ◽  
Acyl Carrier Protein ◽  
Acid Synthesis ◽  
Simultaneous Increase ◽  
Carrier Protein ◽  
Carrier Proteins ◽  
Acyl Carrier Proteins ◽  
Malonyl Coa

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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