Analysis of the Yarrowia lipolytica proteome reveals subtle variations in expression levels between lipogenic and non-lipogenic conditions

2021 ◽  
Vol 21 (2) ◽  
Author(s):  
Ryan Sestric ◽  
Vic Spicer ◽  
Oleg V Krokhin ◽  
Richard Sparling ◽  
David B Levin
Keyword(s):  
Yarrowia Lipolytica ◽  
Carbon Flux ◽  
Neutral Lipids ◽  
Pyruvate Decarboxylase ◽  
Atp Citrate Lyase ◽  
Expression Levels ◽  
Rich Media ◽  
Citrate Lyase ◽  
The Relationship

ABSTRACT Oleaginous yeasts have the ability to store greater than 20% of their mass as neutral lipids, in the form of triacylglycerides. The ATP citrate lyase is thought to play a key role in triacylglyceride synthesis, but the relationship between expression levels of this and other related enzymes is not well understood in the role of total lipid accumulation conferring the oleaginous phenotype. We conducted comparative proteomic analyses with the oleaginous yeast, Yarrowia lipolytica, grown in either nitrogen-sufficient rich media or nitrogen-limited minimal media. Total proteins extracted from cells collected during logarithmic and late stationary growth phases were analyzed by 1D liquid chromatography, followed by mass spectroscopy. The ATP citrate lyase enzyme was expressed at similar concentrations in both conditions, in both logarithmic and stationary phase, but many upstream and downstream enzymes showed drastically different expression levels. In non-lipogenic conditions, several pyruvate enzymes were expressed at higher concentration. These enzymes, especially the pyruvate decarboxylase and pyruvate dehydrogenase, may be regulating carbon flux away from central metabolism and reducing the amount of citrate being produced in the mitochondria. While crucial for the oleaginous phenotype, the constitutively expressed ATP citrate lyase appears to cleave citrate in response to carbon flux upstream from other enzymes creating the oleaginous phenotype.

scholarly journals Comparative Metabolites and Citrate-Degrading Enzymes Activities in Citrus Fruits Reveal the Role of Balance between ACL and Cyt-ACO in Metabolite Conversions

Plants ◽  
2020 ◽  
Vol 9 (3) ◽  
pp. 350 ◽  
Author(s):  
Lingxia Guo ◽  
Yongzhong Liu ◽  
Lijuan Luo ◽  
Syed Bilal Hussain ◽  
Yingxin Bai ◽  
...  
Keyword(s):  
Carbon Flux ◽  
Acid Metabolism ◽  
Soluble Sugars ◽  
Citrus Fruits ◽  
Atp Citrate Lyase ◽  
Acetyl Coa ◽  
Degrading Enzymes ◽  
Citrate Lyase ◽  
Fruit Development And Ripening

Citric acid metabolism is considered to be the central cellular process of metabolite conversions. ATP-citrate lyase (ACL) and cytosolic aconitase (cyt-ACO) are the two citrate-degrading enzymes that decide the carbon flux towards different metabolite biosynthesis pathways. However, the correlation of their activities with metabolite concentrations in citrus fruits is still unclear. Here, the concentrations of soluble sugars, organic acids, acetyl-CoA, flavonoids, carotenoids, and γ-aminobutyric acid, as well as the activities of ACL, cyt-ACO, acetyl-CoA C-acetyltransferase, and acetyl-CoA carboxylase, were compared among the fruits of six citrus cultivars during fruit development and ripening. The results showed that the correlation between citrate concentration and cyt-ACO or ACL activity varied greatly among cultivars, while the activities of cyt-ACO and ACL had a significantly negative correlation (r = −0.4431). Moreover, ACL overexpression and RNA interference in the Citrus callus indicated that increasing and decreasing the ACL activity could reduce and induce cyt-ACO activity, respectively. In addition, significant correlation was only observed between the ACL activity and the concentration of acetyl-CoA (r = 0.4333). Taken together, the present study suggested that ACL and cyt-ACO synergistically control the citrate fate for the biosynthesis of other metabolites, but they are not the key determinants for the accumulation of citrate, as well as other metabolites in citrus fruits.

The role of ATP citrate lyase in the transfer of acetyl groups in rat liver

1968 ◽  
Vol 158 (1) ◽  
pp. 51-61 ◽  
Author(s):  
Yasushi Daikuhara ◽  
Takuo Tsunemi ◽  
Yoshiro Takeda
Keyword(s):  
Rat Liver ◽  
Atp Citrate Lyase ◽  
Citrate Lyase

scholarly journals Engineering of the Pyruvate Dehydrogenase Bypass inSaccharomyces cerevisiae: Role of the Cytosolic Mg2+ and Mitochondrial K+ Acetaldehyde Dehydrogenases Ald6p and Ald4p in Acetate Formation during Alcoholic Fermentation

2000 ◽  
Vol 66 (8) ◽  
pp. 3151-3159 ◽  
Author(s):  
Fabienne Remize ◽  
Emilie Andrieu ◽  
Sylvie Dequin
Keyword(s):  
Carbon Flux ◽  
Alcoholic Fermentation ◽  
Pyruvate Decarboxylase ◽  
Wine Yeast ◽  
Wild Type ◽  
Acetate Production ◽  
Wine Yeast Strain ◽  
Acetate Formation ◽  
The Impact

ABSTRACT Acetic acid plays a crucial role in the organoleptic balance of many fermented products. We have investigated the factors controlling the production of acetate by Saccharomyces cerevisiaeduring alcoholic fermentation by metabolic engineering of the enzymatic steps involved in its formation and its utilization. The impact of reduced pyruvate decarboxylase (PDC), limited acetaldehyde dehydrogenase (ACDH), or increased acetoacetyl coenzyme A synthetase (ACS) levels in a strain derived from a wine yeast strain was studied during alcoholic fermentation. In the strain with the PDC1gene deleted exhibiting 25% of the PDC activity of the wild type, no significant differences were observed in the acetate yield or in the amounts of secondary metabolites formed. A strain overexpressingACS2 and displaying a four- to sevenfold increase in ACS activity did not produce reduced acetate levels. In contrast, strains with one or two disrupted copies of ALD6, encoding the cytosolic Mg2+-activated NADP-dependent ACDH and exhibiting 60 and 30% of wild-type ACDH activity, showed a substantial decrease in acetate yield (the acetate production was 75 and 40% of wild-type production, respectively). This decrease was associated with a rerouting of carbon flux towards the formation of glycerol, succinate, and butanediol. The deletion of ALD4, encoding the mitochondrial K+-activated NAD(P)-linked ACDH, had no effect on the amount of acetate formed. In contrast, a strain lacking both Ald6p and Ald4p exhibited a long delay in growth and acetate production, suggesting that Ald4p can partially replace the Ald6p isoform. Moreover, the ald6 ald4 double mutant was still able to ferment large amounts of sugar and to produce acetate, suggesting the contribution of another member(s) of the ALDfamily.

Molecular biology of acetyl-CoA metabolism

2000 ◽  
Vol 28 (6) ◽  
pp. 591-593 ◽  
Author(s):  
B. J. Nikolau ◽  
D. J. Oliver ◽  
P. S. Schnable ◽  
E. S. Wurtele
Keyword(s):  
Pyruvate Dehydrogenase ◽  
Pyruvate Dehydrogenase Complex ◽  
Pyruvate Decarboxylase ◽  
Acetyl Coa Carboxylase ◽  
Atp Citrate Lyase ◽  
Acetyl Coa ◽  
Acetaldehyde Dehydrogenase ◽  
Citrate Lyase ◽  
Biochemical Genetic ◽  
Dehydrogenase Complex

We have characterized the expression of potential acetyl-CoA-generating genes (acetyl-CoA synthetase, pyruvate decarboxylase, acetaldehyde dehydrogenase, plastidic pyruvate dehydrogenase complex and ATP-citrate lyase), and compared these with the expression of acetyl-CoA-metabolizing genes (heteromeric and homomeric acetyl-CoA carboxylase). These comparisons have led to the development of testable hypotheses as to how distinct pools of acetyl-CoA are generated and metabolized. These hypotheses are being tested by combined biochemical, genetic and molecular biological experiments, which is providing insights into how acetyl-CoA metabolism is regulated.

scholarly journals NOCTURNIN Gene Diurnal Variation in Healthy Volunteers and Expression Levels in Shift Workers

2019 ◽  
Vol 2019 ◽  
pp. 1-8 ◽  
Author(s):  
Massimo Bracci ◽  
Alfredo Copertaro ◽  
Veronica Ciarapica ◽  
Mariella Barbaresi ◽  
Stefano Esposito ◽  
...  
Keyword(s):  
Diurnal Variation ◽  
Healthy Volunteers ◽  
Peripheral Blood ◽  
Human Peripheral Blood ◽  
Peripheral Blood Lymphocytes ◽  
Shift Workers ◽  
Expression Levels ◽  
Blood Lymphocytes ◽  
The Relationship

Objective. The NOCTURNIN gene links nutrient absorption and metabolism to the circadian clock. Shift workers are at a heightened risk of overweight and of developing obesity and metabolic syndrome. This study investigates the diurnal variation of NOCTURNIN in healthy volunteers and its expression levels in rotational shift and daytime workers. Methods. NOCTURNIN expression levels were evaluated in peripheral blood lymphocytes from 15 healthy volunteers at 4-hour intervals for 24 h. Metabolic parameters and NOCTURNIN expression were measured in workers engaged in shift and daytime work. Results. In the group of volunteers NOCTURNIN expression showed diurnal variation, with a peak at 8:00 AM. NOCTURNIN expression was higher in shift workers than in daytime workers. Multivariate analysis confirmed the role of shift work as an independent factor affecting NOCTURNIN expression. Notably, its level correlated directly with body mass index and inversely with total energy expenditure. Conclusions. Measuring NOCTURNIN expression levels in human peripheral blood lymphocytes can improve investigations on the relationship between changes in circadian rhythm and metabolic disorders. Shift workers show higher NOCTURNIN levels than daytime workers.

Role of ATP in the ATP citrate lyase reaction

1967 ◽  
Vol 26 (5) ◽  
pp. 602-608 ◽  
Author(s):  
Hideo Inoue ◽  
Fujio Suzuki ◽  
Hiroaki Tanioka ◽  
Yoshiro Takeda
Keyword(s):  
Atp Citrate Lyase ◽  
Citrate Lyase

scholarly journals Both Subunits of ATP-Citrate Lyase from Chlorobium tepidum Contribute to Catalytic Activity

2006 ◽  
Vol 188 (18) ◽  
pp. 6544-6552 ◽  
Author(s):  
Wonduck Kim ◽  
F. Robert Tabita
Keyword(s):  
Catalytic Mechanism ◽  
Specific Activity ◽  
High Specific Activity ◽  
Small Subunit ◽  
High Yield ◽  
Chlorobium Tepidum ◽  
Atp Citrate Lyase ◽  
Citrate Lyase ◽  
Green Sulfur

ABSTRACT ATP-citrate lyase (ACL) is an essential enzyme of the reductive tricarboxylic acid (RTCA) pathway of CO2 assimilation. The RTCA pathway occurs in several groups of autotrophic prokaryotes, including the green sulfur bacteria. ACL catalyzes the coenzyme A (CoA)-dependent and MgATP-dependent cleavage of citrate into oxaloacetate and acetyl-CoA, representing a key step in the RTCA pathway. To characterize this enzyme from the green sulfur bacterium Chlorobium tepidum and determine the role of its two distinct polypeptide chains, recombinant holo-ACL as well as its two individual subunit polypeptides were synthesized in Escherichia coli. The recombinant holoenzyme, prepared from coexpressed large and small ACL genes, and the individual large and small subunit polypeptides, prepared from singly expressed genes, were all purified to homogeneity to high yield. Purified recombinant holo-ACL was isolated at high specific activity, and its k cat was comparable to that of previously prepared native C. tepidum ACL. Moreover, the purified recombinant large and small subunit polypeptides were able to reconstitute the holo-ACL in vitro, with activity levels approaching that of recombinant holo-ACL prepared from coexpressed genes. Stoichiometric amounts of each subunit protein were required to maximize the activity and form the most stable structure of reconstituted holo-ACL. These results suggested that this reconstitution system could be used to discern the catalytic role of specific amino acid residues on each subunit. Reconstitution and mutagenesis studies together indicated that residues of each subunit contributed to different aspects of the catalytic mechanism, suggesting that both subunit proteins contribute to the active site of C. tepidum ACL.

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