Limits...
Critical Involvement of Environmental Carbon Dioxide Fixation to Drive Wax Ester Fermentation in Euglena

View Article: PubMed Central - PubMed

ABSTRACT

Accumulation profiles of wax esters in Euglena gracilis Z were studied under several environmental conditions. The highest amount of total wax esters accumulated under hypoxia in the dark, and C28 (myristyl-myristate, C14:0-C14:0) was prevalent among all conditions investigated. The wax ester production was almost completely suppressed under anoxia in the light, and supplying exogenous inorganic carbon sources restored wax ester fermentation, indicating the need for external carbon sources for the wax ester fermentation. 13C-labeling experiments revealed specific isotopic enrichment in the odd-numbered fatty acids derived from wax esters, indicating that the exogenously-supplied CO2 was incorporated into wax esters via the propionyl-CoA pathway through the reverse tricarboxylic acid (TCA) cycle. The addition of 3-mercaptopicolinic acid, a phosphoenolpyruvate carboxykinase (PEPCK) inhibitor, significantly affected the incorporation of 13C into citrate and malate as the biosynthetic intermediates of the odd-numbered fatty acids, suggesting the involvement of PEPCK reaction to drive wax ester fermentation. Additionally, the 13C-enrichment pattern of succinate suggested that the CO2 assimilation might proceed through alternative pathways in addition to the PEPCK reaction. The current results indicate that the mechanisms of anoxic CO2 assimilation are an important target to reinforce wax ester fermentation in Euglena.

No MeSH data available.


Related in: MedlinePlus

13C-labeling time course of organic acids in the presence or absence of 3MPA.(A) Relative peak areas of organic acids in the presence (black symbol) or absence (white symbol) of 3MPA. (B) Relative abundances of M, M+1 and M+2 ions characteristic to each organic acid in the presence (3MPA-treated cells, black symbol) or absence (control cells, white symbol) of 3MPA. Values and error bars are the mean and standard deviation from triplicate cultures. Asterisks and daggers indicate statistically significant differences. Red (3MPA treatment) and blue (control) indicate statistically significant differences between 0 min and respective time points for each treatment. Student’s t-test with Bonferroni correction; statistical significance between treatments at each time point (green); Student’s t-test; † p < 0.10, *p < 0.05, **p < 0.01, ***p < 0.001.
© Copyright Policy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC5036851&req=5

pone.0162827.g005: 13C-labeling time course of organic acids in the presence or absence of 3MPA.(A) Relative peak areas of organic acids in the presence (black symbol) or absence (white symbol) of 3MPA. (B) Relative abundances of M, M+1 and M+2 ions characteristic to each organic acid in the presence (3MPA-treated cells, black symbol) or absence (control cells, white symbol) of 3MPA. Values and error bars are the mean and standard deviation from triplicate cultures. Asterisks and daggers indicate statistically significant differences. Red (3MPA treatment) and blue (control) indicate statistically significant differences between 0 min and respective time points for each treatment. Student’s t-test with Bonferroni correction; statistical significance between treatments at each time point (green); Student’s t-test; † p < 0.10, *p < 0.05, **p < 0.01, ***p < 0.001.

Mentions: Fig 5 shows the time course experiments to monitor the CO2 assimilation under the anoxic-light condition. Cells in the modified KH medium were supplied with 13CO2 gas after 10 min of N2 gas aeration. Oxaloacetate (OAA) and fumarate were not detectable under our experimental conditions. Under anoxic-light conditions, the citrate levels started to decline within 5 min of supplying 13CO2 and it reached about 57% (without 3MPA, control cells) and 48% (3MPA-treated cells) of the initial level at 60 min, and were consistently higher in the control cells during the period (Fig 5A). The malate levels also decreased and reached about 62% (control) and 41% (3MPA-treated cells) of the initial level during 60 min (Fig 5A). A continuous decrease in the citrate and malate levels (Fig 5A), irrespective of 3MPA treatment, indicated that the pre-accumulated metabolite pools in the normal KH medium were consumed during wax ester fermentation in the modified KH medium. However, succinate levels continued to be elevated during the experiments irrespective of the 3MPA treatments (Fig 5A). The natural isotope (M) abundances (Fig 5B) suggested that the 3MPA treatment inhibited consumption of the pre-accumulated citrate and malate metabolic pools and that succinate metabolism was apparently unaffected by 3MPA.


Critical Involvement of Environmental Carbon Dioxide Fixation to Drive Wax Ester Fermentation in Euglena
13C-labeling time course of organic acids in the presence or absence of 3MPA.(A) Relative peak areas of organic acids in the presence (black symbol) or absence (white symbol) of 3MPA. (B) Relative abundances of M, M+1 and M+2 ions characteristic to each organic acid in the presence (3MPA-treated cells, black symbol) or absence (control cells, white symbol) of 3MPA. Values and error bars are the mean and standard deviation from triplicate cultures. Asterisks and daggers indicate statistically significant differences. Red (3MPA treatment) and blue (control) indicate statistically significant differences between 0 min and respective time points for each treatment. Student’s t-test with Bonferroni correction; statistical significance between treatments at each time point (green); Student’s t-test; † p < 0.10, *p < 0.05, **p < 0.01, ***p < 0.001.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC5036851&req=5

pone.0162827.g005: 13C-labeling time course of organic acids in the presence or absence of 3MPA.(A) Relative peak areas of organic acids in the presence (black symbol) or absence (white symbol) of 3MPA. (B) Relative abundances of M, M+1 and M+2 ions characteristic to each organic acid in the presence (3MPA-treated cells, black symbol) or absence (control cells, white symbol) of 3MPA. Values and error bars are the mean and standard deviation from triplicate cultures. Asterisks and daggers indicate statistically significant differences. Red (3MPA treatment) and blue (control) indicate statistically significant differences between 0 min and respective time points for each treatment. Student’s t-test with Bonferroni correction; statistical significance between treatments at each time point (green); Student’s t-test; † p < 0.10, *p < 0.05, **p < 0.01, ***p < 0.001.
Mentions: Fig 5 shows the time course experiments to monitor the CO2 assimilation under the anoxic-light condition. Cells in the modified KH medium were supplied with 13CO2 gas after 10 min of N2 gas aeration. Oxaloacetate (OAA) and fumarate were not detectable under our experimental conditions. Under anoxic-light conditions, the citrate levels started to decline within 5 min of supplying 13CO2 and it reached about 57% (without 3MPA, control cells) and 48% (3MPA-treated cells) of the initial level at 60 min, and were consistently higher in the control cells during the period (Fig 5A). The malate levels also decreased and reached about 62% (control) and 41% (3MPA-treated cells) of the initial level during 60 min (Fig 5A). A continuous decrease in the citrate and malate levels (Fig 5A), irrespective of 3MPA treatment, indicated that the pre-accumulated metabolite pools in the normal KH medium were consumed during wax ester fermentation in the modified KH medium. However, succinate levels continued to be elevated during the experiments irrespective of the 3MPA treatments (Fig 5A). The natural isotope (M) abundances (Fig 5B) suggested that the 3MPA treatment inhibited consumption of the pre-accumulated citrate and malate metabolic pools and that succinate metabolism was apparently unaffected by 3MPA.

View Article: PubMed Central - PubMed

ABSTRACT

Accumulation profiles of wax esters in Euglena gracilis Z were studied under several environmental conditions. The highest amount of total wax esters accumulated under hypoxia in the dark, and C28 (myristyl-myristate, C14:0-C14:0) was prevalent among all conditions investigated. The wax ester production was almost completely suppressed under anoxia in the light, and supplying exogenous inorganic carbon sources restored wax ester fermentation, indicating the need for external carbon sources for the wax ester fermentation. 13C-labeling experiments revealed specific isotopic enrichment in the odd-numbered fatty acids derived from wax esters, indicating that the exogenously-supplied CO2 was incorporated into wax esters via the propionyl-CoA pathway through the reverse tricarboxylic acid (TCA) cycle. The addition of 3-mercaptopicolinic acid, a phosphoenolpyruvate carboxykinase (PEPCK) inhibitor, significantly affected the incorporation of 13C into citrate and malate as the biosynthetic intermediates of the odd-numbered fatty acids, suggesting the involvement of PEPCK reaction to drive wax ester fermentation. Additionally, the 13C-enrichment pattern of succinate suggested that the CO2 assimilation might proceed through alternative pathways in addition to the PEPCK reaction. The current results indicate that the mechanisms of anoxic CO2 assimilation are an important target to reinforce wax ester fermentation in Euglena.

No MeSH data available.


Related in: MedlinePlus