scholarly journals In vitro kinetic study of the squalestatin tetraketide synthase dehydratase reveals the stereochemical course of a fungal highly reducing polyketide synthase

2017 ◽  
Vol 53 (10) ◽  
pp. 1727-1730 ◽  
Author(s):  
Emma Liddle ◽  
Alan Scott ◽  
Li-Chen Han ◽  
David Ivison ◽  
Thomas J. Simpson ◽  
...  
Keyword(s):  
Fatty Acid ◽  
Kinetic Study ◽  
Fatty Acid Synthase ◽  
Polyketide Synthase ◽  
Substrate Selectivity ◽  
Dh Domain

The substrate selectivity of the isolated dehydratase (DH) domain of a fungal highly-reducing polyketide synthase is closely related to that of mammalian fatty acid synthase.

scholarly journals Structural comparison of Acinetobacter baumannii β-ketoacyl-acyl carrier protein reductases in fatty acid and aryl polyene biosynthesis

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Woo Cheol Lee ◽  
Sungjae Choi ◽  
Ahjin Jang ◽  
Kkabi Son ◽  
Yangmee Kim
Keyword(s):  
Fatty Acid ◽  
Acinetobacter Baumannii ◽  
Gene Cluster ◽  
Fatty Acid Synthase ◽  
Acyl Carrier Protein ◽  
Gene Clusters ◽  
Carrier Protein ◽  
Aryl Group ◽  
Visible Absorption

AbstractSome Gram-negative bacteria harbor lipids with aryl polyene (APE) moieties. Biosynthesis gene clusters (BGCs) for APE biosynthesis exhibit striking similarities with fatty acid synthase (FAS) genes. Despite their broad distribution among pathogenic and symbiotic bacteria, the detailed roles of the metabolic products of APE gene clusters are unclear. Here, we determined the crystal structures of the β-ketoacyl-acyl carrier protein (ACP) reductase ApeQ produced by an APE gene cluster from clinically isolated virulent Acinetobacter baumannii in two states (bound and unbound to NADPH). An in vitro visible absorption spectrum assay of the APE polyene moiety revealed that the β-ketoacyl-ACP reductase FabG from the A. baumannii FAS gene cluster cannot be substituted for ApeQ in APE biosynthesis. Comparison with the FabG structure exhibited distinct surface electrostatic potential profiles for ApeQ, suggesting a positively charged arginine patch as the cognate ACP-binding site. Binding modeling for the aryl group predicted that Leu185 (Phe183 in FabG) in ApeQ is responsible for 4-benzoyl moiety recognition. Isothermal titration and arginine patch mutagenesis experiments corroborated these results. These structure–function insights of a unique reductase in the APE BGC in comparison with FAS provide new directions for elucidating host–pathogen interaction mechanisms and novel antibiotics discovery.

scholarly journals A Fatty Acid Synthase Gene in Cochliobolus carbonum Required for Production of HC-Toxin, Cyclo(d-Prolyl-l-Alanyl- d-Alanyl- l-2-Amino-9,10-Epoxi-8-Oxodecanoyl)

1997 ◽  
Vol 10 (2) ◽  
pp. 207-214 ◽  
Author(s):  
Joong-Hoon Ahn ◽  
Jonathan D. Walton
Keyword(s):  
Fatty Acid ◽  
Fatty Acid Synthase ◽  
Cyclic Peptide ◽  
Decanoic Acid ◽  
Toxin Production ◽  
Gene Encoding ◽  
Cochliobolus Carbonum ◽  
Primary Amino ◽  
Hc Toxin

The fungal maize pathogen Cochliobolus carbonum produces a phytotoxic and cytostatic cyclic peptide, HC-toxin, of structure cyclo(D-prolyl-L-alanyl-D-alanyl-L-Aeo), in which Aeo stands for 2-amino-9,10-epoxi-8-oxodecanoic acid. Here we report the isolation of a gene, TOXC, that is present only in HC-toxin-producing (Tox2+) fungal strains. TOXC is present in most Tox2+ strains in three functional copies, all of which are on the same chromosome as the gene encoding HC-toxin synthetase. When all copies of TOXC are mutated by targeted gene disruption, the fungus grows and sporulates normally in vitro but no longer makes HC-toxin and is not pathogenic, indicating that TOXC has a specific role in HC-toxin production and hence virulence. The TOXC mRNA is 6.5 kb and the predicted product has 2,080 amino acids and a molecular weight of 233,000. The primary amino acid sequence is highly similar (45 to 47% identity) to the β subunit of fatty acid synthase from several lower eukaryotes, and contains, in the same order as in other β subunits, domains predicted to encode acetyl transferase, enoyl reductase, dehydratase, and malonyl-palmityl transferase. The most plausible function of TOXC is to contribute to the synthesis of the decanoic acid backbone of Aeo.

scholarly journals Targeting de novo lipogenesis and the Lands cycle induces ferroptosis in KRAS-mutant lung cancer

2021 ◽  
Author(s):  
Caterina Bartolacci ◽  
Cristina Andreani ◽  
Goncalo Dias do Vale ◽  
Stefano Berto ◽  
Margherita Melegari ◽  
...  
Keyword(s):  
Lung Cancer ◽  
Fatty Acid ◽  
Metabolic Networks ◽  
Fatty Acid Synthase ◽  
De Novo ◽  
Genetically Engineered ◽  
De Novo Lipogenesis ◽  
Main Process

Mutant KRAS (KM) is the most common oncogene in lung cancer (LC). KM regulates several metabolic networks, but their role in tumorigenesis is still not sufficiently characterized to be exploited in cancer therapy. To identify metabolic networks specifically deregulated in KMLC, we characterized the lipidome of genetically engineered LC mice, cell lines, patient derived xenografts and primary human samples. We also determined that KMLC, but not EGFR-mutant (EGFR-MUT) LC, is enriched in triacylglycerides (TAG) and phosphatidylcholines (PC). We also found that KM upregulates fatty acid synthase (FASN), a rate-limiting enzyme in fatty acid (FA) synthesis promoting the synthesis of palmitate and PC. We determined that FASN is specifically required for the viability of KMLC, but not of LC harboring EGFR-MUT or wild type KRAS. Functional experiments revealed that FASN inhibition leads to ferroptosis, a reactive oxygen species (ROS)-and iron-dependent cell death. Consistently, lipidomic analysis demonstrated that FASN inhibition in KMLC leads to accumulation of PC with polyunsaturated FA (PUFA) chains, which are the substrate of ferroptosis. Integrating lipidomic, transcriptome and functional analyses, we demonstrated that FASN provides saturated (SFA) and monounsaturated FA (MUFA) that feed the Lands cycle, the main process remodeling oxidized phospholipids (PL), such as PC. Accordingly, either inhibition of FASN or suppression of the Lands cycle enzymes PLA2 and LPCAT3, promotes the intracellular accumulation of lipid peroxides and ferroptosis in KMLC both in vitro and in vivo. Our work supports a model whereby the high oxidative stress caused by KM dictates a dependency on newly synthesized FA to repair oxidated phospholipids, establishing a targetable vulnerability. These results connect KM oncogenic signaling, FASN induction and ferroptosis, indicating that FASN inhibitors already in clinical trial in KMLC patients (NCT03808558) may be rapidly deployed as therapy for KMLC.

scholarly journals A kinetic rationale for functional redundancy in fatty acid biosynthesis

2020 ◽  
Vol 117 (38) ◽  
pp. 23557-23564
Author(s):  
Alex Ruppe ◽  
Kathryn Mains ◽  
Jerome M. Fox
Keyword(s):  
Fatty Acids ◽  
Fatty Acid ◽  
Fatty Acid Synthase ◽  
Unsaturated Fatty Acids ◽  
Fatty Acid Biosynthesis ◽  
Average Length ◽  
Functional Redundancy ◽  
Experimental Investigations ◽  
Total Production

Cells build fatty acids with biocatalytic assembly lines in which a subset of enzymes often exhibit overlapping activities (e.g., two enzymes catalyze one or more identical reactions). Although the discrete enzymes that make up fatty acid pathways are well characterized, the importance of catalytic overlap between them is poorly understood. We developed a detailed kinetic model of the fatty acid synthase (FAS) ofEscherichia coliand paired that model with a fully reconstituted in vitro system to examine the capabilities afforded by functional redundancy in fatty acid synthesis. The model captures—and helps explain—the effects of experimental perturbations to FAS systems and provides a powerful tool for guiding experimental investigations of fatty acid assembly. Compositional analyses carried out in silico and in vitro indicate that FASs with multiple partially redundant enzymes enable tighter (i.e., more independent and/or broader range) control of distinct biochemical objectives—the total production, unsaturated fraction, and average length of fatty acids—than FASs with only a single multifunctional version of each enzyme (i.e., one enzyme with the catalytic capabilities of two partially redundant enzymes). Maximal production of unsaturated fatty acids, for example, requires a second dehydratase that is not essential for their synthesis. This work provides a kinetic, control-theoretic rationale for the inclusion of partially redundant enzymes in fatty acid pathways and supplies a valuable framework for carrying out detailed studies of FAS kinetics.

scholarly journals Genome Sequencing and Analysis of Thraustochytriidae sp. SZU445 Provides Novel Insights into the Polyunsaturated Fatty Acid Biosynthesis Pathway

Marine Drugs ◽  
2020 ◽  
Vol 18 (2) ◽  
pp. 118 ◽  
Author(s):  
Xingyu Zhu ◽  
Shuangfei Li ◽  
Liangxu Liu ◽  
Siting Li ◽  
Yanqing Luo ◽  
...  
Keyword(s):  
Fatty Acid ◽  
Fatty Acid Synthase ◽  
Polyketide Synthase ◽  
Fatty Acid Biosynthesis ◽  
Docosapentaenoic Acid ◽  
Omega 3 ◽  
Gene Ontology Database ◽  
Fatty Acid Biosynthesis Pathway ◽  
Fas Pathway ◽  
Acid Pathway

Thraustochytriidae sp. have broadly gained attention as a prospective resource for the production of omega-3 fatty acids production in significant quantities. In this study, the whole genome of Thraustochytriidae sp. SZU445, which produces high levels of docosapentaenoic acid (DPA) and docosahexaenoic acid (DHA), was sequenced and subjected to protein annotation. The obtained clean reads (63.55 Mb in total) were assembled into 54 contigs and 25 scaffolds, with maximum and minimum lengths of 400 and 0.0054 Mb, respectively. A total of 3513 genes (24.84%) were identified, which could be classified into six pathways and 44 pathway groups, of which 68 genes (1.93%) were involved in lipid metabolism. In the Gene Ontology database, 22,436 genes were annotated as cellular component (8579 genes, 38.24%), molecular function (5236 genes, 23.34%), and biological process (8621 genes, 38.42%). Four enzymes corresponding to the classic fatty acid synthase (FAS) pathway and three enzymes corresponding to the classic polyketide synthase (PKS) pathway were identified in Thraustochytriidae sp. SZU445. Although PKS pathway-associated dehydratase and isomerase enzymes were not detected in Thraustochytriidae sp. SZU445, a putative DHA- and DPA-specific fatty acid pathway was identified.

scholarly journals Fisetin Protects Against Hepatic Steatosis Through Regulation of the Sirt1/AMPK and Fatty Acid β-Oxidation Signaling Pathway in High-Fat Diet-Induced Obese Mice

2018 ◽  
Vol 49 (5) ◽  
pp. 1870-1884 ◽  
Author(s):  
Chian-Jiun Liou ◽  
Ciao-Han Wei ◽  
Ya-Ling Chen ◽  
Ching-Yi Cheng ◽  
Chia-Ling Wang ◽  
...  
Keyword(s):  
Fatty Acid ◽  
Lipid Metabolism ◽  
Hepatic Steatosis ◽  
Lipid Accumulation ◽  
High Fat Diet ◽  
Fatty Acid Synthase ◽  
Epididymal Adipose Tissue ◽  
Obese Mice ◽  
High Fat

Background/Aims: Fisetin is a naturally abundant flavonoid isolated from various fruits and vegetables that was recently identified to have potential biological functions in improving allergic airway inflammation, as well as anti-oxidative and anti-tumor properties. Fisetin has also been demonstrated to have anti-obesity properties in mice. However, the effect of fisetin on nonalcoholic fatty liver disease (NAFLD) is still elusive. Thus, the present study evaluated whether fisetin improves hepatic steatosis in high-fat diet (HFD)-induced obese mice and regulates lipid metabolism of FL83B hepatocytes in vitro. Methods: NAFLD was induced by HFD in male C57BL/6 mice. The mice were then injected intraperitoneally with fisetin for 10 weeks. In another experiment, FL83B cells were challenged with oleic acid to induce lipid accumulation and treated with various concentrations of fisetin. Results: NAFLD mice treated with fisetin had decreased body weight and epididymal adipose tissue weight compared to NAFLD mice. Fisetin treatment also reduced liver lipid droplet and hepatocyte steatosis, alleviated serum free fatty acid, and leptin concentrations, significantly decreased fatty acid synthase, and significantly increased phosphorylation of AMPKα and the production of sirt-1 and carnitine palmitoyltransferase I in the liver tissue. In vitro, fisetin decreased lipid accumulation and increased lipolysis and β-oxidation in hepatocytes. Conclusion: This study suggests that fisetin is a potential novel treatment for alleviating hepatic lipid metabolism and improving NAFLD in mice via activation of the sirt1/AMPK and β-oxidation pathway.

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