Biotin carboxyl carrier protein isoforms in Brassicaceae oilseeds

2000 ◽  
Vol 28 (6) ◽  
pp. 595-598 ◽  
Author(s):  
J. J. Thelen ◽  
S. Mekhedov ◽  
J. B. Ohlrogge
Keyword(s):  
Fatty Acid Biosynthesis ◽  
De Novo ◽  
Single Gene ◽  
Protein Isoforms ◽  
Carrier Protein ◽  
Biotin Carboxyl Carrier Protein ◽  
Developing Seeds ◽  
Malonyl Coa ◽  
Evolutionarily Conserved ◽  
Plant Arabidopsis Thaliana

De novo fatty acid biosynthesis occurs predominantly in plastids. The committed step for this pathway is the production of malonyl-CoA catalysed by acetyl-CoA carboxylase (ACCase). In most plants, plastidial ACCase is a multisubunit complex minimally comprised of four polypeptides, which catalyse two reactions. In the simple oilseed plant, Arabidopsis thaliana two cDNAs encoding biotin carboxyl carrier protein (BCCP) isoforms have been identified. The remaining three subunits of ACCase appear to be single gene members in A. thaliana [Mekhedov, Martinez de Ilarduya and Ohlrogge (2000) Plant Physiol. 122, 389–401]. Transcript and protein analyses indicate that BCCP isoform 1 is constitutively expressed while isoform 2 is predominantly expressed in developing seeds. The apparent masses of constitutive and seed-enriched BCCP isoforms agree with the apparent masses of recombinantly expressed isoforms 1 and 2, respectively. In a related oilseed, Brassica napus multiple putative BCCP polypeptides were also observed in developing seeds. The presence of a divergent class of BCCP genes in A. thaliana and B. napus coincident with appropriately sized biotin-containing proteins expressed specifically in developing seeds, suggests that these BCCPs play an evolutionarily conserved role in oil deposition.

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.

Use of 13C in biosynthetic studies. The labeling pattern in dihydrofusarubin enriched from [13C]- and [13C, 2H]acetate in cultures of Fusarium solani

1980 ◽  
Vol 58 (13) ◽  
pp. 1380-1385 ◽  
Author(s):  
Itsuo Kurobane ◽  
Leo C. Vining ◽  
A. Gavin McInnes ◽  
John A. Walter
Keyword(s):  
Fusarium Solani ◽  
Carrier Protein ◽  
Single Chain ◽  
Acetyl Coa ◽  
Biotin Carboxyl Carrier Protein ◽  
H Nmr ◽  
Malonyl Coa ◽  
Starter Unit ◽  
Biosynthetic Studies ◽  
Labeling Pattern

The pattern of 13C and 2H incorporation from [1-13C]-, [2-13C]-, [1-13C0;1;1,2-13C1;0;1]-, and [2-13C0;1,2-2H3;3]acetate into dihydrofusarubin 1, produced by cultures of Fusariumsolani, has been determined by 13C and 2H nmr of the derivatives anhydrofusarubin 3 and anhydrofusarubin diacetate 4. The results show that 1 is biosynthesized from seven uniformly-incorporated acetate units with C-3, C-11 originating from the "starter" unit. They strongly suggest that linear head-to-tail condensation of an acetate and six malonate units forms a single-chain heptaketide intermediate. The evidence also suggests that, during conversion of [13C, 2H]-labeled acetyl-CoA to malonyl-CoA, 2H is transferred to biotin carboxyl carrier protein where it does not exchange rapidly with the medium and is available for conversion of endogenous malonyl-CoA to [2H]-enriched acetyl-CoA.

scholarly journals Elucidation of transient protein-protein interactions within carrier protein-dependent biosynthesis

2020 ◽  
Author(s):  
Michael Burkart ◽  
Thomas Bartholow ◽  
Terra Sztain ◽  
Ashay Patel ◽  
D Lee ◽  
...  
Keyword(s):  
Fatty Acid ◽  
Protein Interactions ◽  
Fatty Acid Biosynthesis ◽  
De Novo ◽  
Acyl Carrier Protein ◽  
Protein Docking ◽  
Carrier Protein ◽  
Protein Protein Interactions ◽  
Acid Biosynthesis ◽  
E Coli

Abstract Fatty acid biosynthesis (FAB) is an essential and highly conserved metabolic pathway. In bacteria, this process is mediated by an elaborate network of protein•protein interactions (PPIs) involving a small, dynamic acyl carrier protein that interacts with dozens of other partner proteins (PPs). These PPIs have remained poorly characterized due to their dynamic and transient nature. Using a combination of solution-phase NMR spectroscopy and protein-protein docking simulations, we report a comprehensive residue-by-residue comparison of the PPIs formed during FAB in Escherichia coli. This work reveals the molecular basis of six discrete binding events responsible for E. coli FAB and offers insights into a method to characterize these events and those in related carrier protein-dependent pathways. ONE SENTENCE SUMMARY: Through a combination of structural and computational analysis, a comparative evaluation of protein-protein interactions in de novo fatty acid biosynthesis in E. coli is performed.

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 A Soybean Acyl Carrier Protein, GmACP, Is Important for Root Nodule Symbiosis

2014 ◽  
Vol 27 (5) ◽  
pp. 415-423 ◽  
Author(s):  
Jun Wang ◽  
Katalin Tóth ◽  
Kiwamu Tanaka ◽  
Cuong T. Nguyen ◽  
Zhe Yan ◽  
...  
Keyword(s):  
Fatty Acid ◽  
Fatty Acid Biosynthesis ◽  
Fluorescent Protein ◽  
De Novo ◽  
Acyl Carrier Protein ◽  
Fatty Acid Content ◽  
Gas Chromatography Mass Spectrometry ◽  
Carrier Protein ◽  
Acid Biosynthesis ◽  
Transgenic Roots

Legumes (members of family Fabaceae) establish a symbiotic relationship with nitrogen-fixing soil bacteria (rhizobia) to overcome nitrogen source limitation. Single root hair epidermal cells serve as the entry point for bacteria to infect the host root, leading to development of a new organ, the nodule, which the bacteria colonize. In the present study, the putative role of a soybean acyl carrier protein (ACP), GmACP (Glyma18g47950), was examined in nodulation. ACP represent an essential cofactor protein in fatty acid biosynthesis. Phylogenetic analysis of plant ACP protein sequences showed that GmACP was classified in a legume-specific clade. Quantitative reverse-transcription polymerase chain reaction analysis demonstrated that GmACP was expressed in all soybean tissues but showed higher transcript accumulation in nodule tissue. RNA interference-mediated gene silencing of GmACP resulted in a significant reduction in nodule numbers on soybean transgenic roots. Fluorescent protein-labeled GmACP was localized to plastids in planta, the site of de novo fatty acid biosynthesis in plants. Analysis of the fatty acid content of root tissue silenced for GmACP expression, as determined by gas chromatography–mass spectrometry, showed an approximately 22% reduction, specifically in palmitic and stearic acid. Taken together, our data provide evidence that GmACP plays an important role in nodulation.

scholarly journals Elongation of fatty acids by microsomal fractions from the brain of the developing rat

1975 ◽  
Vol 152 (3) ◽  
pp. 495-501 ◽  
Author(s):  
P J Brophy ◽  
D E Vance
Keyword(s):  
Fatty Acids ◽  
Fatty Acid ◽  
Rat Brain ◽  
Microsomal Fraction ◽  
Fatty Acid Biosynthesis ◽  
De Novo ◽  
Soluble Fraction ◽  
Residual Activity ◽  
Acid Biosynthesis ◽  
Malonyl Coa

Elongation of fatty acids by microsomal fractions obtained from rat brain was measured by the incorporation of [2-14C]malonyl-CoA into fatty in the presence of palmitoyl-CoA or stearoyl-CoA. 2. Soluble and microsomal fractions were prepared from 21-day-old rats; density gradient centrifugation demonstrated that the stearoyl-CoA elongation system was localized in the microsomal fraction whereas fatty acid biosynthesis de novo from acetyl-CoA occurred in the soluble fraction. The residual activity de novo in the microsomal fraction was attributed to minor contamination by the soluble fraction. 3. The optimum concentration of [2-14C]malonyl-CoA for elongation of fatty acids was 25 mum for palmitoyl-CoA or stearoyl-CoA, and the corresponding optimum concentrations for the two primer acyl-CoA esters were 8.0 and 7.2 muM respectively. 4. Nadph was the preferred cofactor for fatty acid formation from palmitoyl-CoA or stearoyl-CoA, although NADH could partially replace it. 5. The stearoyl-CoA elongation system required a potassium phosphate buffer concentration of 0.075M for maximum activity; CoA (1 MUM) inhibited this elongation system by approx. 30%. 6. The fatty acids formed from malonyl-CoA and palmitoyl-CoA had a predominant chain length of C18 whereas stearoyl-CoA elongation resulted in an even distribution of fatty acids with chain lengths of C20, C22 and C24. 7. The products of stearoyl-CoA elongation were identified as primarily unesterified fatty acids. 8. The developmental pattern of fatty acid biosynthesis by rat brain microsomal preparations was studied and both the palmitoyl-CoA and stearoyl-CoA elongation systems showed large increases in activity between days 10 and 18 after birth.

Studies on the Inhibition of Biotin-Containing Carboxylases by Acetyl-CoA Carboxylase Inhibitors

1993 ◽  
Vol 48 (3-4) ◽  
pp. 294-300 ◽  
Author(s):  
Anja Motel ◽  
Simone Günther ◽  
Martin Clauss ◽  
Klaus Kobek ◽  
Manfred Focke ◽  
...  
Keyword(s):  
Fatty Acid ◽  
Fatty Acid Biosynthesis ◽  
De Novo ◽  
Higher Plants ◽  
Acetate Incorporation ◽  
Acetyl Coa Carboxylase ◽  
Acid Biosynthesis ◽  
Acetyl Coa ◽  
Malonyl Coa ◽  
Key Enzyme

In higher plants the biosynthetic machinery of de novo fatty acid biosynthesis, measured as [14C]acetate incorporation into fatty acids, is predominantly located in plastids. A key enzyme in this pathway is the biotin-containing acetyl-CoA carboxylase (ACC , EC 6.4.1.2) which catalyzes the ATP-dependent carboxylation of acetyl-CoA to malonyl-CoA. The ACC from Poaceae is very efficiently blocked by two herbicide classes, the cyclohexane-1,3-diones (e.g. sethoxydim, cycloxydim) and the aryloxyphenoxy-propionic acids (e.g. diclofop, fluazifop). It is shown that within the Poaceae not only different species but also different varieties exist which exhibit an altered sensitivity and tolerance towards both herbicide classes, which points to a mutation of the target enzyme ACC. In purifying the ACC we extended our research to the possible presence of other biotin-containing plant enzymes. In protein preparations from maize, oat, barley, pea and lentil we were able to demonstrate the carboxylation of acetyl-CoA, propionyl-CoA and methylcrotonyl-CoA. The two herbicide classes not only block the ACC, but also the activity of the propionyl-CoA carboxylase (PCC ), whereas the methylcrotonyl- CoA carboxylase (MCC ), a distinct biotin-containing enzyme from mitochondria, is not affected. MCC may play a role in isoprenoid catabolism. Whether PCC is a separate plastid enzyme or only a side activity of ACC is under current investigation. The efficiency of the graminicides in sensitive Poaceae is then not only determined by the inhibition of ACC, malonyl-CoA and fatty acid biosynthesis, but also by the exclusion of the PCC-catalyzed metabolic pathways of the plant cell.

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