Characterization of Helianthus annuus Lipoic Acid Biosynthesis: The Mitochondrial Octanoyltransferase and Lipoyl Synthase Enzyme System

Lipoic acid (LA, 6,8-dithiooctanoic acid) is a sulfur containing coenzyme essential for the activity of several key enzymes involved in oxidative and single carbon metabolism in most bacteria and eukaryotes. LA is synthetized by the concerted activity of the octanoyltransferase (LIP2, EC 2.3.1.181) and lipoyl synthase (LIP1, EC 2.8.1.8) enzymes. In plants, pyruvate dehydrogenase (PDH), 2-oxoglutarate dehydrogenase or glycine decarboxylase are essential complexes that need to be lipoylated. These lipoylated enzymes and complexes are located in the mitochondria, while PDH is also present in plastids where it provides acetyl-CoA for de novo fatty acid biosynthesis. As such, lipoylation of PDH could regulate fatty acid synthesis in both these organelles. In the present work, the sunflower LIP1 and LIP2 genes (HaLIP1m and HaLIP2m) were isolated sequenced, cloned, and characterized, evaluating their putative mitochondrial location. The expression of these genes was studied in different tissues and protein docking was modeled. The genes were also expressed in Escherichia coli and Arabidopsis thaliana, where their impact on fatty acid and glycerolipid composition was assessed. Lipidomic studies in Arabidopsis revealed lipid remodeling in lines overexpressing these enzymes and the involvement of both sunflower proteins in the phenotypes observed is discussed in the light of the results obtained.

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Function of Heterologous Mycobacterium tuberculosis InhA, a Type 2 Fatty Acid Synthase Enzyme Involved in Extending C20 Fatty Acids to C60-to-C90 Mycolic Acids, during De Novo Lipoic Acid Synthesis in Saccharomyces cerevisiae

Applied and Environmental Microbiology ◽

10.1128/aem.00655-08 ◽

2008 ◽

Vol 74 (16) ◽

pp. 5078-5085 ◽

Cited By ~ 13

Author(s):

Aner Gurvitz ◽

J. Kalervo Hiltunen ◽

Alexander J. Kastaniotis

Keyword(s):

Fatty Acids ◽

Fatty Acid ◽

Lipoic Acid ◽

Fatty Acid Synthase ◽

Fatty Acid Biosynthesis ◽

De Novo ◽

Acid Synthesis ◽

Acid Biosynthesis ◽

Mycolic Acids

ABSTRACT We describe the physiological function of heterologously expressed Mycobacterium tuberculosis InhA during de novo lipoic acid synthesis in yeast (Saccharomyces cerevisiae) mitochondria. InhA, representing 2-trans-enoyl-acyl carrier protein reductase and the target for the front-line antituberculous drug isoniazid, is involved in the activity of dissociative type 2 fatty acid synthase (FASII) that extends associative type 1 fatty acid synthase (FASI)-derived C20 fatty acids to form C60-to-C90 mycolic acids. Mycolic acids are major constituents of the protective layer around the pathogen that contribute to virulence and resistance to certain antimicrobials. Unlike FASI, FASII is thought to be incapable of de novo biosynthesis of fatty acids. Here, the genes for InhA (Rv1484) and four similar proteins (Rv0927c, Rv3485c, Rv3530c, and Rv3559c) were expressed in S. cerevisiae etr1Δ cells lacking mitochondrial 2-trans-enoyl-thioester reductase activity. The phenotype of the yeast mutants includes the inability to produce sufficient levels of lipoic acid, form mitochondrial cytochromes, respire, or grow on nonfermentable carbon sources. Yeast etr1Δ cells expressing mitochondrial InhA were able to respire, grow on glycerol, and produce lipoic acid. Commensurate with a role in mitochondrial de novo fatty acid biosynthesis, InhA could accept in vivo much shorter acyl-thioesters (C4 to C8) than was previously thought (>C12). Moreover, InhA functioned in the absence of AcpM or protein-protein interactions with its native FASII partners KasA, KasB, FabD, and FabH. None of the four proteins similar to InhA complemented the yeast mutant phenotype. We discuss the implications of our findings with reference to lipoic acid synthesis in M. tuberculosis and the potential use of yeast FASII mutants for investigating the physiological function of drug-targeted pathogen enzymes involved in fatty acid biosynthesis.

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

10.21203/rs.3.rs-72626/v1 ◽

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.

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Inhibition of early steps of de novo fatty-acid biosynthesis by different xenobiotica

Physiologia Plantarum ◽

10.1034/j.1399-3054.1991.810217.x ◽

1991 ◽

Vol 81 (2) ◽

pp. 251-255

Author(s):

Manfred Focke ◽

Andrea Feld ◽

Hartmut K. Lichtenthaler

Keyword(s):

Fatty Acid ◽

Fatty Acid Biosynthesis ◽

De Novo ◽

Acid Biosynthesis

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Genome-Wide Mapping of Histone H3 Lysine 4 Trimethylation (H3K4me3) and Its Involvement in Fatty Acid Biosynthesis in Sunflower Developing Seeds

Plants ◽

10.3390/plants10040706 ◽

2021 ◽

Vol 10 (4) ◽

pp. 706

Author(s):

Antonio J. Moreno-Pérez ◽

Raquel Martins-Noguerol ◽

Cristina DeAndrés-Gil ◽

Mónica Venegas-Calerón ◽

Rosario Sánchez ◽

...

Keyword(s):

Fatty Acid ◽

Chromatin Remodeling ◽

Molecular Mechanisms ◽

Acid Metabolism ◽

Fatty Acid Biosynthesis ◽

Acid Synthesis ◽

Sunflower Seeds ◽

Acid Biosynthesis ◽

Functional Regions ◽

Genome Wide

Histone modifications are of paramount importance during plant development. Investigating chromatin remodeling in developing oilseeds sheds light on the molecular mechanisms controlling fatty acid metabolism and facilitates the identification of new functional regions in oil crop genomes. The present study characterizes the epigenetic modifications H3K4me3 in relationship with the expression of fatty acid-related genes and transcription factors in developing sunflower seeds. Two master transcriptional regulators identified in this analysis, VIV1 (homologous to Arabidopsis ABI3) and FUS3, cooperate in the regulation of WRINKLED 1, a transcriptional factor regulating glycolysis, and fatty acid synthesis in developing oilseeds.

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Inhibition of de novo Fatty-Acid Biosynthesis in Isolated Chloroplasts by the Antibiotic Cerulenin and Its Synthetic Derivatives

Zeitschrift für Naturforschung C ◽

10.1515/znc-1992-0611 ◽

1992 ◽

Vol 47 (5-6) ◽

pp. 382-386 ◽

Cited By ~ 6

Author(s):

Bernd List ◽

Andrea Golz ◽

Wilhelm Boland ◽

Hartmut K. Lichtenthaler

Keyword(s):

Fatty Acid ◽

Inhibitory Activity ◽

Fatty Acid Biosynthesis ◽

De Novo ◽

Specific Inhibitor ◽

Hydrocarbon Chain ◽

Acetate Incorporation ◽

Acid Biosynthesis ◽

Structural Element ◽

Isolated Chloroplasts

The antibiotic cerulenin was shown to be a potent dose-dependent inhibitor of de novo fattyacid biosynthesis in intact isolated chloroplasts of different plants (measured as [14C]acetate incorporation into the total fatty-acid fraction). Various chemical derivatives of cerulenin were synthesized and tested in the chloroplast assay-system of oat, spinach and pea. Modifications of the hydrocarbon chain of cerulenin (e.g. tetrahydro-cerulenin and its short-chain cis-2,3-epoxy-4-oxoheptanamide derivative) decreased the inhibitory activity of cerulenin, whereas variations of the epoxy-oxo-amide structural element led to a complete loss of inhibition potency. The results indicate that the naturally occurring antibiotic cerulenin is the most active specific inhibitor of de novo fatty-acid biosynthesis, but the formation of the hydroxylactam ring seems to be an essential requirement for the inhibitory activity. Those structural analogues of cerulenin, which can no longer form a hydroxylactam ring, do not possess any inhibitory capacity.

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SYNTHESIS OF FATTY ACIDS BY TESTICULAR TISSUE IN VITRO

Canadian Journal of Biochemistry and Physiology ◽

10.1139/y63-142 ◽

1963 ◽

Vol 41 (1) ◽

pp. 1267-1274

Author(s):

Peter F. Hall ◽

Edward E. Nishizawa ◽

Kristen B. Eik-Nes

Keyword(s):

Fatty Acids ◽

Fatty Acid ◽

Fatty Acid Biosynthesis ◽

De Novo ◽

Fatty Acid Fraction ◽

Testicular Tissue ◽

Acid Biosynthesis ◽

Adrenal Tissue ◽

Substrate Fatty Acid

The fatty acids palmitic, palmitoleic, stearic, and oleic have been isolated from rabbit testis and evidence for the synthesis of palmitic and stearic acids de novo from acetate-1-C14is presented. ICSH did not produce demonstrable stimulation of the synthesis of these acids in vitro although the hormone stimulated the production of testosterone-C14by the same tissue. Adrenal tissue was shown to contain palmitic, stearic, and oleic acids, and ACTH did not increase the incorporation of acetate-1-C14into a fatty acid fraction extracted following incubation of adrenal tissue in the presence of this substrate. Fatty acid biosynthesis, therefore, is probably not influenced by the mechanisms by which tropic hormones increase steroid formation.

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Effect of Thiolactomycin on de novo Fatty Acid Biosynthesis in Plants

Zeitschrift für Naturforschung C ◽

10.1515/znc-1990-0537 ◽

1990 ◽

Vol 45 (5) ◽

pp. 518-520 ◽

Cited By ~ 4

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.

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De novo Fatty Acid Biosynthesis Contributes Significantly to Establishment of a Bioenergetically Favorable Environment for Vaccinia Virus Infection

PLoS Pathogens ◽

10.1371/journal.ppat.1004021 ◽

2014 ◽

Vol 10 (3) ◽

pp. e1004021 ◽

Cited By ~ 54

Author(s):

Matthew D. Greseth ◽

Paula Traktman

Keyword(s):

Fatty Acid ◽

Virus Infection ◽

Vaccinia Virus ◽

Fatty Acid Biosynthesis ◽

De Novo ◽

Acid Biosynthesis ◽

Vaccinia Virus Infection ◽

Favorable Environment

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Reconstruction of the Fatty Acid Biosynthetic Pathway ofExiguobacterium antarcticumB7 Based on Genomic and Bibliomic Data

BioMed Research International ◽

10.1155/2016/7863706 ◽

2016 ◽

Vol 2016 ◽

pp. 1-9 ◽

Cited By ~ 2

Author(s):

Regiane Kawasaki ◽

Rafael A. Baraúna ◽

Artur Silva ◽

Marta S. P. Carepo ◽

Rui Oliveira ◽

...

Keyword(s):

Fatty Acids ◽

Fatty Acid ◽

Fatty Acid Biosynthesis ◽

De Novo ◽

Short Chain Fatty Acids ◽

Hexadecenoic Acid ◽

Biosynthesis Pathway ◽

Acid Biosynthesis ◽

Cold Environments ◽

Fatty Acid Biosynthesis Pathway

Exiguobacterium antarcticumB7 is extremophile Gram-positive bacteria able to survive in cold environments. A key factor to understanding cold adaptation processes is related to the modification of fatty acids composing the cell membranes of psychrotrophic bacteria. In our study we show thein silicoreconstruction of the fatty acid biosynthesis pathway ofE. antarcticumB7. To build the stoichiometric model, a semiautomatic procedure was applied, which integrates genome information using KEGG and RAST/SEED. Constraint-based methods, namely, Flux Balance Analysis (FBA) and elementary modes (EM), were applied. FBA was implemented in the sense of hexadecenoic acid production maximization. To evaluate the influence of the gene expression in the fluxome analysis, FBA was also calculated using thelog2⁡FCvalues obtained in the transcriptome analysis at 0°C and 37°C. The fatty acid biosynthesis pathway showed a total of 13 elementary flux modes, four of which showed routes for the production of hexadecenoic acid. The reconstructed pathway demonstrated the capacity ofE. antarcticumB7 tode novoproduce fatty acid molecules. Under the influence of the transcriptome, the fluxome was altered, promoting the production of short-chain fatty acids. The calculated models contribute to better understanding of the bacterial adaptation at cold environments.

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