Phosphoenolpyruvate carboxylase identified as a key enzyme in erythrocytic Plasmodium falciparum carbon metabolism

Janet Storm, Sonal Sethia, Gavin Blackburn, A Chokkathukalam, David Watson, R. Breitling, Graham Coombs, S. Muller

Research output: Contribution to journalArticle

24 Citations (Scopus)

Abstract

Phospoenolpyruvate carboxylase (PEPC) is absent from humans but encoded in the Plasmodium falciparum genome, suggesting that PEPC has a parasite-specific function. To investigate its importance in P. falciparum, we generated a pepc null mutant (D10Δpepc), which was only achievable when malate, a reduction product of oxaloacetate, was added to the growth medium. D10Δpepc had a severe growth defect in vitro, which was partially reversed by addition of malate or fumarate, suggesting that pepc may be essential in vivo. Targeted metabolomics using 13C-U-D-glucose and 13C-bicarbonate showed that the conversion of glycolytically-derived PEP into malate, fumarate, aspartate and citrate was abolished in D10Δpepc and that pentose phosphate pathway metabolites and glycerol 3-phosphate were present at increased levels. In contrast, metabolism of the carbon skeleton of 13C,15N-U-glutamine was similar in both parasite lines, although the flux was lower in D10Δpepc; it also confirmed the operation of a complete forward TCA cycle in the wild type parasite. Overall, these data confirm the CO2 fixing activity of PEPC and suggest that it provides metabolites essential for TCA cycle anaplerosis and the maintenance of cytosolic and mitochondrial redox balance. Moreover, these findings imply that PEPC may be an exploitable target for future drug discovery.
LanguageEnglish
Article numbere1003876
Pages1-14
Number of pages14
JournalPLOS Pathogens
Volume10
Issue number1
DOIs
Publication statusPublished - 16 Jan 2014

Fingerprint

Phosphoenolpyruvate Carboxylase
Plasmodium falciparum
Fumarates
Parasites
Carbon
Enzymes
Oxaloacetic Acid
Pentose Phosphate Pathway
Metabolomics
Drug Discovery
Bicarbonates
Growth
Glutamine
Skeleton
Aspartic Acid
Citric Acid
Oxidation-Reduction
Maintenance
Genome
Glucose

Keywords

  • phosphoenolpyruvate carboxylase
  • key enzyme
  • metabolism
  • erythrocytic
  • falciparum
  • plasmodium
  • carbon

Cite this

Storm, J., Sethia, S., Blackburn, G., Chokkathukalam, A., Watson, D., Breitling, R., ... Muller, S. (2014). Phosphoenolpyruvate carboxylase identified as a key enzyme in erythrocytic Plasmodium falciparum carbon metabolism. PLOS Pathogens, 10(1), 1-14. [e1003876]. https://doi.org/10.1371/journal.ppat.1003876
Storm, Janet ; Sethia, Sonal ; Blackburn, Gavin ; Chokkathukalam, A ; Watson, David ; Breitling, R. ; Coombs, Graham ; Muller, S. / Phosphoenolpyruvate carboxylase identified as a key enzyme in erythrocytic Plasmodium falciparum carbon metabolism. In: PLOS Pathogens. 2014 ; Vol. 10, No. 1. pp. 1-14.
@article{20ca0401bada4defa962f4ae5110d110,
title = "Phosphoenolpyruvate carboxylase identified as a key enzyme in erythrocytic Plasmodium falciparum carbon metabolism",
abstract = "Phospoenolpyruvate carboxylase (PEPC) is absent from humans but encoded in the Plasmodium falciparum genome, suggesting that PEPC has a parasite-specific function. To investigate its importance in P. falciparum, we generated a pepc null mutant (D10Δpepc), which was only achievable when malate, a reduction product of oxaloacetate, was added to the growth medium. D10Δpepc had a severe growth defect in vitro, which was partially reversed by addition of malate or fumarate, suggesting that pepc may be essential in vivo. Targeted metabolomics using 13C-U-D-glucose and 13C-bicarbonate showed that the conversion of glycolytically-derived PEP into malate, fumarate, aspartate and citrate was abolished in D10Δpepc and that pentose phosphate pathway metabolites and glycerol 3-phosphate were present at increased levels. In contrast, metabolism of the carbon skeleton of 13C,15N-U-glutamine was similar in both parasite lines, although the flux was lower in D10Δpepc; it also confirmed the operation of a complete forward TCA cycle in the wild type parasite. Overall, these data confirm the CO2 fixing activity of PEPC and suggest that it provides metabolites essential for TCA cycle anaplerosis and the maintenance of cytosolic and mitochondrial redox balance. Moreover, these findings imply that PEPC may be an exploitable target for future drug discovery.",
keywords = "phosphoenolpyruvate carboxylase, key enzyme, metabolism, erythrocytic, falciparum, plasmodium, carbon",
author = "Janet Storm and Sonal Sethia and Gavin Blackburn and A Chokkathukalam and David Watson and R. Breitling and Graham Coombs and S. Muller",
year = "2014",
month = "1",
day = "16",
doi = "10.1371/journal.ppat.1003876",
language = "English",
volume = "10",
pages = "1--14",
journal = "PLOS Pathogens",
issn = "1553-7366",
number = "1",

}

Phosphoenolpyruvate carboxylase identified as a key enzyme in erythrocytic Plasmodium falciparum carbon metabolism. / Storm, Janet; Sethia, Sonal; Blackburn, Gavin; Chokkathukalam, A; Watson, David; Breitling, R.; Coombs, Graham; Muller, S.

In: PLOS Pathogens, Vol. 10, No. 1, e1003876, 16.01.2014, p. 1-14.

Research output: Contribution to journalArticle

TY - JOUR

T1 - Phosphoenolpyruvate carboxylase identified as a key enzyme in erythrocytic Plasmodium falciparum carbon metabolism

AU - Storm, Janet

AU - Sethia, Sonal

AU - Blackburn, Gavin

AU - Chokkathukalam, A

AU - Watson, David

AU - Breitling, R.

AU - Coombs, Graham

AU - Muller, S.

PY - 2014/1/16

Y1 - 2014/1/16

N2 - Phospoenolpyruvate carboxylase (PEPC) is absent from humans but encoded in the Plasmodium falciparum genome, suggesting that PEPC has a parasite-specific function. To investigate its importance in P. falciparum, we generated a pepc null mutant (D10Δpepc), which was only achievable when malate, a reduction product of oxaloacetate, was added to the growth medium. D10Δpepc had a severe growth defect in vitro, which was partially reversed by addition of malate or fumarate, suggesting that pepc may be essential in vivo. Targeted metabolomics using 13C-U-D-glucose and 13C-bicarbonate showed that the conversion of glycolytically-derived PEP into malate, fumarate, aspartate and citrate was abolished in D10Δpepc and that pentose phosphate pathway metabolites and glycerol 3-phosphate were present at increased levels. In contrast, metabolism of the carbon skeleton of 13C,15N-U-glutamine was similar in both parasite lines, although the flux was lower in D10Δpepc; it also confirmed the operation of a complete forward TCA cycle in the wild type parasite. Overall, these data confirm the CO2 fixing activity of PEPC and suggest that it provides metabolites essential for TCA cycle anaplerosis and the maintenance of cytosolic and mitochondrial redox balance. Moreover, these findings imply that PEPC may be an exploitable target for future drug discovery.

AB - Phospoenolpyruvate carboxylase (PEPC) is absent from humans but encoded in the Plasmodium falciparum genome, suggesting that PEPC has a parasite-specific function. To investigate its importance in P. falciparum, we generated a pepc null mutant (D10Δpepc), which was only achievable when malate, a reduction product of oxaloacetate, was added to the growth medium. D10Δpepc had a severe growth defect in vitro, which was partially reversed by addition of malate or fumarate, suggesting that pepc may be essential in vivo. Targeted metabolomics using 13C-U-D-glucose and 13C-bicarbonate showed that the conversion of glycolytically-derived PEP into malate, fumarate, aspartate and citrate was abolished in D10Δpepc and that pentose phosphate pathway metabolites and glycerol 3-phosphate were present at increased levels. In contrast, metabolism of the carbon skeleton of 13C,15N-U-glutamine was similar in both parasite lines, although the flux was lower in D10Δpepc; it also confirmed the operation of a complete forward TCA cycle in the wild type parasite. Overall, these data confirm the CO2 fixing activity of PEPC and suggest that it provides metabolites essential for TCA cycle anaplerosis and the maintenance of cytosolic and mitochondrial redox balance. Moreover, these findings imply that PEPC may be an exploitable target for future drug discovery.

KW - phosphoenolpyruvate carboxylase

KW - key enzyme

KW - metabolism

KW - erythrocytic

KW - falciparum

KW - plasmodium

KW - carbon

UR - http://www.plospathogens.org/

U2 - 10.1371/journal.ppat.1003876

DO - 10.1371/journal.ppat.1003876

M3 - Article

VL - 10

SP - 1

EP - 14

JO - PLOS Pathogens

T2 - PLOS Pathogens

JF - PLOS Pathogens

SN - 1553-7366

IS - 1

M1 - e1003876

ER -