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

1975 ◽  
Vol 146 (2) ◽  
pp. 439-445 ◽  
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.

Effect of Thiolactomycin on de novo Fatty Acid Biosynthesis in Plants

1990 ◽  
Vol 45 (5) ◽  
pp. 518-520 ◽  
Author(s):  
Manfred Focke ◽  
Andrea Feld ◽  
Hartmut K. Lichtenthaler
Keyword(s):  
Fatty Acids ◽  
Fatty Acid ◽  
Fatty Acid Biosynthesis ◽  
De Novo ◽  
Fatty Acid Synthetase ◽  
Acetate Incorporation ◽  
Acid Biosynthesis ◽  
Acetyl Coa ◽  
Malonyl Coa ◽  
Total Fatty Acids

Thiolactomycin was shown to be a potent inhibitor of de novo fatty acid biosynthesis in intact isolated chloroplasts (measured as [14C]acetate incorporation into total fatty acids). In our attempt to further localize the inhibition site we confirmed the inhibition with a fatty acid synthetase preparation, measuring the incorporation of [14C]malonyl-CoA into total fatty acids. From the two proposed enzymic targets of the fatty acid synthetase by thiolactomycin we could exclude the acetyl-CoA: ACP transacetylase. It appears that the inhibition by thiolactomycin occurs on the level of the condensing enzymes, i.e. the 3-oxoacyl-ACP synthases. We also demonstrated that the two starting enzymes of de novo fatty acid biosynthesis, the acetyl-CoA synthetase and the acetyl-CoA carboxylase, are not affected by thiolactomycin.

Fatty Acid Synthetase from Pythium debaryanum

1973 ◽  
Vol 51 (3) ◽  
pp. 241-248 ◽  
Author(s):  
Simon W. T. Law ◽  
David N. Burton
Keyword(s):  
Fatty Acid ◽  
De Novo ◽  
Acyl Carrier Protein ◽  
Fatty Acid Synthetase ◽  
Apparent Size ◽  
Supernatant Fraction ◽  
Acetyl Coa ◽  
Ph Optimum ◽  
Malonyl Coa ◽  
Long Chain

The presence of a fatty acid synthetase, capable of de novo synthesis of long chain fatty acids from acetyl-CoA, malonyl-CoA, and NADPH has been demonstrated in the simple fungus Pythium debaryanum (ATCC 9998), a member of the class Oomycetes. The enzyme is found in the supernatant fraction after centrifugation at 123 000 g for 35 min.The fatty acid synthetase was demonstrated to be of the multienzyme complex type, showing no dependence on acyl carrier protein for activity. The fatty acid products were identified as principally a mixture of free and esterified palmitic and stearic acids by gas-radiochromatography, and were shown to be synthesized de novo by the ratio of incorporation of acetyl- and malonyl-CoA, the Schmidt decarboxylation, and the lack of activity of medium and long chain acyl-CoA derivatives as acceptors of two carbon units in an elongation reaction. The pH optimum for the reaction was 6.8, and the Km for acetyl-CoA was 3.3 μM.The molecular weight of the fatty acid synthetase was estimated by gel filtration on Sepharose-4B as being of the order of 4 × 106. Attempts to reduce the apparent size of the enzyme by treatment with detergents and various enzymes were unsuccessful.

On the purification of a fatty acid synthetase complex from an insect

1976 ◽  
Vol 54 (8) ◽  
pp. 1397-1399 ◽  
Author(s):  
S. N. Thompson ◽  
J. S. Barlow
Keyword(s):  
Fatty Acids ◽  
Fatty Acid ◽  
Column Chromatography ◽  
De Novo ◽  
Gel Filtration ◽  
Fatty Acid Synthetase ◽  
De Novo Synthesis ◽  
Anion Exchange Column ◽  
Malonyl Coa ◽  
Salt Fractionation

The fatty acid synthetase complex of the blowfly, Lucilia sericata, catalyzing the de novo synthesis of fatty acids from acetyl- and malonyl-CoA (EC 2.3.1.9 and EC 2.3.1.39 respectively) and requiring NADPH, was isolated from whole-insect homogenates by ultracentrifugation. Purification was carried out by salt fractionation, anion exchange column chromatography on ECTEOLA (epichlorohydrin triethanolamine) cellulose, and gel filtration column chromatography on Sephadex®.

Notes: Inhibition of the Acetyl-CoA Carboxylase of Barley Chloroplasts by Cycloxydim and Sethoxydim

1987 ◽  
Vol 42 (11-12) ◽  
pp. 1361-1363 ◽  
Author(s):  
Manfred Focke ◽  
Hartmut K. Lichtenthaler
Keyword(s):  
Fatty Acids ◽  
Fatty Acid ◽  
Enzyme Preparation ◽  
Fatty Acid Biosynthesis ◽  
De Novo ◽  
Acetyl Coa Carboxylase ◽  
Acid Biosynthesis ◽  
Hordeum Vulgare L ◽  
Acetyl Coa ◽  
Malonyl Coa

The effect of the three cyclohexane-1,3-dione derivatives cycloxydim, sethoxydim and clethodim on the incorpora­tion of 14C-labelled acetate, malonate. acctyl-CoA or malonyl-CoA into fatty acids was studied in an enzyme preparation isolated from barley chloroplasts (Hordeum vulgare L. var. “Alexis”). The herbicides cycloxydim, clethodim and sethoxydim block the de novo fatty acid biosynthesis from [2-14C]acetatc and [1-14C]acetyl-CoA, whereas that of [2-14C]malonatc and [2-14C)malonyl-CoA is not affected. The data indicate that the mode of action of the cyclohexane-1,3-dione derivatives in the sensitive bar­ley plant consists in the inhibition of de novo fatty acid biosynthesis by blocking the acetyl-CoA carboxylase (EC 6.4.1.2.).

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.

scholarly journals Effect of phenylpyruvate on enzymes involved in fatty acid synthesis in rat brain

1973 ◽  
Vol 134 (2) ◽  
pp. 545-555 ◽  
Author(s):  
John M. Land ◽  
John B. Clark
Keyword(s):  
Fatty Acid ◽  
Citrate Synthase ◽  
Mitochondrial Protein ◽  
Fatty Acid Synthetase ◽  
Acetyl Coa Carboxylase ◽  
Acetyl Coa ◽  
Adult Rats ◽  
Malonyl Coa ◽  
Old Rats ◽  
The Brain

1. The activities of, and the effects of phenylpyruvate on, citrate synthase (EC 4.1.3.7), acetyl-CoA carboxylase (EC 6.4.1.2) and fatty acid synthetase derived from the brains of 14-day-old and adult rats were investigated. 2. The brain citrate synthase from 14-day-old rats had a Km for oxaloacetate of 2.38μm and for acetyl-CoA of 16.9μm, and a Vmax. of 838nmol of acetyl-CoA incorporation/min per mg of mitochondrial protein. From adult rat brain this enzyme had a Km for oxaloacetate of 2.5μm and for acetyl-CoA of 16.6μm and a Vmax. of 1070nmol of acetyl-CoA incorporated/min per mg of mitochondrial protein. Phenylpyruvate inhibited the enzyme from adult and young rat brains in a competitive fashion with respect to acetyl-CoA, with a Ki of 700μm. 3. The brain acetyl-CoA carboxylase from 14-day-old rats had a Km for acetyl-CoA of 21μm and a Vmax. of 0.248nmol/min per mg of protein, and from adult rats a Km for acetyl-CoA of 21μm and a Vmax. of 0.173nmol/min per mg of protein. The enzyme from young and adult rats required citrate (Ka=3mm) for activation and were inhibited non-competitively by phenylpyruvate, with a Ki of 10mm. 4. The brain fatty acid synthetase from 14-day-old rats had a Km for acetyl-CoA of 7.58μm and a Vmax. of 1.1 nmol of malonyl-CoA incorporated/min per mg of protein, and from adult rats a Km for acetyl-CoA of 4.9μm and a Vmax. of 0.48nmol of malonyl-CoA incorporated/min per mg of protein. Phenylpyruvate acted as a competitive inhibitor with respect to acetyl-CoA with a Ki of 250μm for the enzyme from 14-day-old rats. 5. These results are discussed with respect to phenylketonuria, and it is suggested that the inhibition of the brain fatty acid synthetase and possibly the citrate synthetase by phenylpyruvate could explain the defective myelination characteristic of this condition.

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