scholarly journals Dynamism of Metabolic Carbon Flow of Starch and Lipids in Chlamydomonas debaryana

2021 ◽  
Vol 12 ◽  
Author(s):  
Naoki Sato ◽  
Masakazu Toyoshima
Keyword(s):  
Fatty Acids ◽  
Growth Phase ◽  
De Novo ◽  
Membrane Lipids ◽  
Carbon Flow ◽  
Added Value ◽  
Gas Chromatography Mass Spectrometry ◽  
Exponential Growth Phase ◽  
Alternative Source ◽  
Metabolic Carbon

Microalgae have the potential to recycle CO2 as starch and triacylglycerol (TAG), which provide alternative source of biofuel and high added-value chemicals. Starch accumulates in the chloroplast, whereas TAG accumulates in the cytoplasmic lipid droplets (LD). Preferential accumulation of starch or TAG may be achieved by switching intracellular metabolic carbon flow, but our knowledge on this control remains limited. Are these two products mutually exclusive? Or, does starch act as a precursor to TAG synthesis, or vice versa? To answer these questions, we analyzed carbon flow in starch and lipids using a stable isotope 13C in Chlamydomonas debaryana NIES-2212, which accumulates, without nutrient limitation, starch in the exponential growth phase and TAG in the stationary phase. Pulse labeling experiments as well as pulse labeling and chase experiments were conducted, and then, gas chromatography-mass spectrometry (GC-MS) analysis was performed on starch-derived glucose and lipid-bound fatty acids. We exploited the previously developed method of isotopomer analysis to estimate the proportion of various pools with different isotopic abundance. Starch turned over rapidly to provide carbon for the synthesis of fatty acids in the exponential phase cells. Most fatty acids showed rapid and slow components of metabolism, whereas oleic acid decayed according to a single exponential curve. Highly labeled population of fatty acids that accumulated during the initial labeling decreased rapidly, and replaced by low abundance population during the chase time, indicating that highly labeled fatty acids were degraded and the resulting carbons were re-used in the re-synthesis with about 9-fold unlabeled, newly fixed carbons. Elongation of C16–C18 acids in vivo was indicated by partially labeled C18 acids. The accumulation of TAG in the stationary growth phase was accounted for by both de novo synthesis and remodeling of membrane lipids. These results suggest that de novo synthesis of starch and TAG was rapid and transient, and also almost independent to each other, but there is a pool of starch quickly turning over for the synthesis of fatty acids. Fatty acids were also subject to re-synthesis. Evidence was also provided for remodeling of lipids, namely, re-use of acyl groups in polar lipids for TAG synthesis.

scholarly journals Characterization of fatty acid desaturases reveals stress-induced synthesis of C18 unsaturated fatty acids enriched in triacylglycerol in the oleaginous alga Chromochloris zofingiensis

2021 ◽  
Vol 14 (1) ◽  
Author(s):  
Tao Wu ◽  
Lihua Yu ◽  
Yu Zhang ◽  
Jin Liu
Keyword(s):  
Mass Spectrometry ◽  
Fatty Acids ◽  
Unsaturated Fatty Acids ◽  
De Novo ◽  
Membrane Lipids ◽  
Membrane Lipid ◽  
Gas Chromatography Mass Spectrometry ◽  
Transcription Start Sites ◽  
Chromatography Mass Spectrometry ◽  
Chromochloris Zofingiensis

Abstract Background The green microalga Chromochloris zofingiensis is capable of producing high levels of triacylglycerol rich in C18 unsaturated fatty acids (UFAs). FA desaturation degree is regulated by FA desaturases (FADs). Nevertheless, it remains largely unknown regarding what FADs are involved in FA desaturations and how these FADs collaborate to contribute to the high abundance of C18 UFAs in triacylglycerol in C. zofingiensis. Results To address these issues, we firstly determined the transcription start sites of 11 putative membrane-bound FAD-coding genes (CzFADs) and updated their gene models. Functional validation of these CzFADs in yeast and cyanobacterial cells revealed that seven are bona fide FAD enzymes with distinct substrates. Combining the validated functions and predicted subcellular compartments of CzFADs and the FA profiles of C. zofingiensis, the FA desaturation pathways in this alga were reconstructed. Furthermore, a multifaceted lipidomic analysis by systematically integrating thin-layer chromatography, gas chromatography–mass spectrometry and liquid chromatography–mass spectrometry techniques was conducted, unraveling profiles of polar membrane lipids in C. zofingiensis and major desaturation steps occurring in these lipids. By correlating transcriptional patterns of CzFAD genes and changes of lipids upon abiotic stress conditions, our results highlighted collaboration of CzFADs for C18 UFA synthesis and supported that both de novo FA synthesis and membrane lipid remodeling contributed C18 UFAs to triacylglycerol for storage. Conclusions Taken together, our study for the first time elucidated the pathways of C18 FA desaturations and comprehensive profiles of polar membrane lipids in C. zofingiensis and shed light on collaboration of CzFADs for the synthesis and enrichment of C18 UFAs in triacylglycerol.

scholarly journals 13,16-Dimethyl Octacosanedioic Acid (iso-Diabolic Acid), a Common Membrane-Spanning Lipid of Acidobacteria Subdivisions 1 and 3

2011 ◽  
Vol 77 (12) ◽  
pp. 4147-4154 ◽  
Author(s):  
Jaap S. Sinninghe Damsté ◽  
W. Irene C. Rijpstra ◽  
Ellen C. Hopmans ◽  
Johan W. H. Weijers ◽  
Bärbel U. Foesel ◽  
...  
Keyword(s):  
Fatty Acids ◽  
High Performance ◽  
Membrane Lipids ◽  
Direct Analysis ◽  
Polar Lipids ◽  
Gas Chromatography Mass Spectrometry ◽  
Content Type ◽  
Alkyl Chains ◽  
Intact Polar Lipids ◽  
Membrane Spanning

ABSTRACTThe distribution of membrane lipids of 17 different strains representing 13 species of subdivisions 1 and 3 of the phylumAcidobacteria, a highly diverse phylum of theBacteria, were examined by hydrolysis and gas chromatography-mass spectrometry (MS) and by high-performance liquid chromatography-MS of intact polar lipids. Upon both acid and base hydrolyses of total cell material, the uncommon membrane-spanning lipid 13,16-dimethyl octacosanedioic acid (iso-diabolic acid) was released in substantial amounts (22 to 43% of the total fatty acids) from all of the acidobacteria studied. This lipid has previously been encountered only in thermophilicThermoanaerobacterspecies but bears a structural resemblance to the alkyl chains of bacterial glycerol dialkyl glycerol tetraethers (GDGTs) that occur ubiquitously in peat and soil and are suspected to be produced by acidobacteria. As reported previously, most species also containediso-C15and C16:1ω7Cas major fatty acids but the presence ofiso-diabolic acid was unnoticed in previous studies, most probably because the complex lipid that contained this moiety was not extractable from the cells; it could only be released by hydrolysis. Direct analysis of intact polar lipids in the Bligh-Dyer extract of three acidobacterial strains, indeed, did not reveal any membrane-spanning lipids containingiso-diabolic acid. In 3 of the 17 strains, ether-boundiso-diabolic acid was detected after hydrolysis of the cells, including one branched GDGT containingiso-diabolic acid-derived alkyl chains. Since the GDGT distribution in soils is much more complex, branched GDGTs in soil likely also originate from other (acido)bacteria capable of biosynthesizing these components.

scholarly journals Evaluation of microalgae polysaccharides as biostimulants of tomato plant defense using metabolomics and biochemical approaches

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Farid Rachidi ◽  
Redouane Benhima ◽  
Yassine Kasmi ◽  
Laila Sbabou ◽  
Hicham El Arroussi
Keyword(s):  
Fatty Acids ◽  
Plant Defense ◽  
Systemic Acquired Resistance ◽  
Membrane Lipids ◽  
Acquired Resistance ◽  
Metabolite Profile ◽  
Gas Chromatography Mass Spectrometry ◽  
Maximum Effect ◽  
Tomato Plants ◽  
Biotic Stresses

AbstractMicroalgal polysaccharides (PSs) may be an effective elicitor agent that can efficiently protect plants against biotic stresses. In this study, wee investigates, the effect of PS obtained from microalgae and cyanobacteria (D. salina MS002, P. tricorontum MS023, Porphyridium sp. MS081, Desmodesmus sp., D. salina MS067 and A. platensis MS001) on the biochemical and metabolomics markers linked to defense pathways in tomato plants. The phenylalanine ammonia lyase (PAL), chitinase, 1,3-beta-glucanase and peroxidase (POX) activities have been improved in tomato plants leaves treated by polysaccharides extracted from P. triocnutum (238.26%); Desmodesmus sp. (19.95%); P. triocnutum (137.50%) and Porphyridium sp. (47.28%) respectively. For proteins, polyphenols and H2O2, the maximum effect was induced by D. salina 067 (55.01%), Porphyridium sp. (3.97%) and A. platensis (35.08%) respectively. On the other hand, Gas Chromatography-mass spectrometry (GC–MS) metabolomics analysis showed that PSs induced the modification of metabolite profile involved in the wax construction of tomato leaves, such as fatty acids, alkanes, alkenes and phytosterol. PS treatments improved the accumulation of fatty acids C16:3, C18:2 and C18:3 released from the membrane lipids as precursors of oxylipin biosynthesis which are signaling molecules of plant defense. In addition, PS treatment induced the accumulation of C18:0 and Azelaic acid which is a regulator of salicylic acid-dependent systemic acquired resistance. However, molecular and metabolic studies can determine more precisely the mode of action of microalgal polysaccharides as biostimulants/elicitors plant defense.

scholarly journals Characterization of key enzymes involved in triacylglycerol biosynthesis in mycobacteria

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Agostina Crotta Asis ◽  
Franco Savoretti ◽  
Matías Cabruja ◽  
Hugo Gramajo ◽  
Gabriela Gago
Keyword(s):  
Phosphatidic Acid ◽  
Growth Phase ◽  
De Novo ◽  
Cell Envelope ◽  
Exponential Growth Phase ◽  
Genes Encoding ◽  
Concomitant Reduction ◽  
Tag Biosynthesis ◽  
Odd Chain Fatty Acids

AbstractPhosphatidic acid phosphatase (PAP) catalyzes the dephosphorylation of phosphatidic acid (PA) yielding diacylglycerol (DAG), the lipid precursor for triacylglycerol (TAG) biosynthesis. PAP activity has a key role in the regulation of PA flux towards TAG or glycerophospholipid synthesis. In this work we have characterized two Mycobacterium smegmatis genes encoding for functional PAP proteins. Disruption of both genes provoked a sharp reduction in de novo TAG biosynthesis in early growth phase cultures under stress conditions. In vivo labeling experiments demonstrated that TAG biosynthesis was restored in the ∆PAP mutant when bacteria reached exponential growth phase, with a concomitant reduction of phospholipid synthesis. In addition, comparative lipidomic analysis showed that the ∆PAP strain had increased levels of odd chain fatty acids esterified into TAGs, suggesting that the absence of PAP activity triggered other rearrangements of lipid metabolism, like phospholipid recycling, in order to maintain the wild type levels of TAG. Finally, the lipid changes observed in the ∆PAP mutant led to defective biofilm formation. Understanding the interaction between TAG synthesis and the lipid composition of mycobacterial cell envelope is a key step to better understand how lipid homeostasis is regulated during Mycobacterium tuberculosis infection.

scholarly journals Measurement of lipogenic flux by deuterium resolved mass spectrometry

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Xiaorong Fu ◽  
Stanisław Deja ◽  
Justin A. Fletcher ◽  
Norma N. Anderson ◽  
Monika Mizerska ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Fatty Acids ◽  
Gas Chromatography ◽  
De Novo ◽  
De Novo Lipogenesis ◽  
Gas Chromatography Mass Spectrometry ◽  
Deuterated Water ◽  
Wide Range ◽  
Serial Measurements ◽  
Insight Into

AbstractDe novo lipogenesis (DNL) is disrupted in a wide range of human disease. Thus, quantification of DNL may provide insight into mechanisms and guide interventions if it can be performed rapidly and noninvasively. DNL flux is commonly measured by 2H incorporation into fatty acids following deuterated water (2H2O) administration. However, the sensitivity of this approach is limited by the natural abundance of 13C, which masks detection of 2H by mass spectrometry. Here we report that high-resolution Orbitrap gas-chromatography mass-spectrometry resolves 2H and 13C fatty acid mass isotopomers, allowing DNL to be quantified using lower 2H2O doses and shorter experimental periods than previously possible. Serial measurements over 24-hrs in mice detects the nocturnal activation of DNL and matches a 3H-water method in mice with genetic activation of DNL. Most importantly, DNL is detected in overnight-fasted humans in less than an hour and is responsive to feeding during a 4-h study. Thus, 2H specific MS provides the ability to study DNL in settings that are currently impractical.

scholarly journals Decreased Triacylglycerol Content and Elevated Contents of Cell Membrane Lipids in Colorectal Cancer Tissue: A Lipidomic Study

2020 ◽  
Vol 9 (4) ◽  
pp. 1095
Author(s):  
Adriana Mika ◽  
Alicja Pakiet ◽  
Aleksandra Czumaj ◽  
Zbigniew Kaczynski ◽  
Ivan Liakh ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Fatty Acids ◽  
Cell Membrane ◽  
Lipid Composition ◽  
Membrane Lipids ◽  
Saturated Fatty Acids ◽  
Gas Chromatography Mass Spectrometry ◽  
Acid Oxidation ◽  
Cancer Tissue ◽  
Cancer Tissues

Recent evidence suggests that lipid composition in cancer tissues may undergo multiple alterations. However, no comprehensive analysis of various lipid groups in colorectal cancer (CRC) tissue has been conducted thus far. To address the problem in question, we determined the contents of triacylglycerols (TG), an energetic substrate, various lipids necessary for cell membrane formation, among them phospholipids (phosphatidylcholine, phosphatidylethanolamine), sphingolipids (sphingomyelin) and cholesterol (free, esterified and total), and fatty acids included in complex lipids. 1H-nuclear magnetic resonance (1H-NMR) and gas chromatography-mass spectrometry (GC-MS) were used to analyze the lipid composition of colon cancer tissue and normal large intestinal mucosa from 25 patients. Compared with normal tissue, cancer tissues had significantly lower TG content, along with elevated levels of phospholipids, sphingomyelin, and cholesterol. Moreover, the content of oleic acid, the main component of TG, was decreased in cancer tissues, whereas the levels of saturated fatty acids and polyunsaturated fatty acids (PUFAs), which are principal components of polar lipids, were elevated. These lipidome rearrangements were associated with the overexpression of genes associated with fatty acid oxidation, and the synthesis of phospholipids and cholesterol. These findings suggest that reprogramming of lipid metabolism might occur in CRC tissue, with a shift towards increased utilization of TG for energy production and enhanced synthesis of membrane lipids, necessary for the rapid proliferation of cancer cells.

scholarly journals Temporal Expression of the Bacillus subtilis secAGene, Encoding a Central Component of the Preprotein Translocase

1999 ◽  
Vol 181 (2) ◽  
pp. 493-500 ◽  
Author(s):  
Markus Herbort ◽  
Michael Klein ◽  
Erik H. Manting ◽  
Arnold J. M. Driessen ◽  
Roland Freudl
Keyword(s):  
Bacillus Subtilis ◽  
Growth Phase ◽  
Exponential Growth ◽  
De Novo ◽  
Stationary Growth Phase ◽  
Extracellular Proteins ◽  
Exponential Growth Phase ◽  
Central Component ◽  
Temporal Expression ◽  
Stationary Growth

ABSTRACT In Bacillus subtilis, the secretion of extracellular proteins strongly increases upon transition from exponential growth to the stationary growth phase. It is not known whether the amounts of some or all components of the protein translocation apparatus are concomitantly increased in relation to the increased export activity. In this study, we analyzed the transcriptional organization and temporal expression of the secA gene, encoding a central component of the B. subtilis preprotein translocase. We found that secA and the downstream gene (prfB) constitute an operon that is transcribed from a vegetative (ςA-dependent) promoter located upstream ofsecA. Furthermore, using different independent methods, we found that secA expression occurred mainly in the exponential growth phase, reaching a maximal value almost precisely at the transition from exponential growth to the stationary growth phase. Following to this maximum, the de novo transcription ofsecA sharply decreased to a low basal level. Since at the time of maximal secA transcription the secretion activity of B. subtilis strongly increases, our results clearly demonstrate that the expression of at least one of the central components of the B. subtilis protein export apparatus is adapted to the increased demand for protein secretion. Possible mechanistic consequences are discussed.

The biochemical composition of Cylindrotheca closterium (Ehrenb.) Rayman net Levin, as a source of biologically active substances in cumulative cultivation

2021 ◽  
pp. 1-9
Author(s):  
Svetlana N. Zheleznova ◽  
Ruslan G. Gevorgiz
Keyword(s):  
Fatty Acids ◽  
Stationary Phase ◽  
Growth Phase ◽  
Dry Weight ◽  
Biologically Active ◽  
Exponential Growth Phase ◽  
Total Lipids ◽  
Active Growth ◽  
Cylindrotheca Closterium ◽  
Dry Biomass

In this work, we studied the biochemical characteristics of the diatom Cylindrotheca closterium in the cumulative mode of cultivation in different phases of growth. According to our data, in the exponential growth phase, C. closterium biomass is characterized by a minimum lipid content of 3% of dry biomass and a maximum carbohydrates content of 23% of dry weight. During the transition of C. closterium culture to the stationary growth phase, as in the case of a representative of diatoms, a rather high accumulation of total lipids is observed – to 28% dry weight, while the carbohydrates content decreases to 7%. At the end of the stationary phase (21st day of cultivation), the content of fatty acids is 50% of total lipids or 12.5% of dry weight. A certain relationship between the concentrations of fucoxanthin, total lipids and polyunsaturated fatty acids (PUFA) in the biomass of C. сlosterium diatom was recorded. During active growth in the exponential phase, the concentration of fucoxanthin was 0.5–1 mg∙g-1 dry weight. During the tranition of the culture to the stationary phase of growth, the concentration of fucoxanthin reached 7±0.2 mg∙g-1 dry biomass. At the end of the stationary phase, the concentration of fucoxanthin reached its maximum value of 20 mg∙g-1 dry weight.

scholarly journals Proteome analysis of xylose metabolism inRhodotorula toruloidesduring lipid production

2019 ◽  
Author(s):  
Ievgeniia A. Tiukova ◽  
Jule Brandenburg ◽  
Johanna Blomqvist ◽  
Sabine Samples ◽  
Nils Mikkelsen ◽  
...  
Keyword(s):  
Fatty Acids ◽  
Fatty Acid ◽  
Carbon Source ◽  
Growth Phase ◽  
Exponential Growth ◽  
Lipid Production ◽  
Proteome Analysis ◽  
Exponential Growth Phase ◽  
Sugar Uptake ◽  
Beta Oxidation

AbstractBackgroundRhodotorula toruloidesis a promising platform organism for production of lipids from lignocellulosic substrates. Little is known about the metabolic aspects of lipid production from the lignocellolosic sugar xylose by oleaginous yeasts in general andR. toruloidesin particular. This study presents the first proteome analysis of the metabolism ofR. toruloidesduring conversion of xylose to lipids.ResultsR. toruloidescultivated on either glucose or xylose was subjected to comparative analysis of its growth dynamics, lipid composition, fatty acid profiles and proteome. The maximum growth and sugar uptake rate of glucose-grownR. toruloidescells were almost twice that of xylose-grown cells. Cultivation on xylose medium resulted in a lower final biomass yield although final cellular lipid content was similar between glucose- and xylose-grown cells. Analysis of lipid classes revealed the presence of monoacylglycerol in the early exponential growth phase as well as a high proportion of free fatty acids. Carbon source-specific changes in lipid profiles were only observed at early exponential growth phase, where C18 fatty acids were more saturated in xylose-grown cells. Proteins involved in sugar transport, initial steps of xylose assimilation and NADPH regeneration were among the proteins whose levels increased the most in xylose-grown cells across all time points. The levels of enzymes involved in the mevalonate pathway, phospholipid biosynthesis and amino acids biosynthesis differed in response to carbon source. In addition, xylose-grown cells contained higher levels of enzymes involved in peroxisomal beta-oxidation and oxidative stress response compared to cells cultivated on glucose.ConclusionsThe results obtained in the present study suggest that sugar import is the limiting step during xylose conversion byR. toruloidesinto lipids. NADPH appeared to be regenerated primarily through pentose phosphate pathway although it may also involve malic enzyme as well as alcohol and aldehyde dehydrogenases. Increases in enzyme levels of both fatty acid biosynthesis and beta-oxidation in xylose-grown cells was predicted to result in a futile cycle. The results presented here are valuable for the development of lipid production processes employingR. toruloideson xylose-containing substrates.

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