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Yuan P, Hendriks EF, Fernandez HR, et al. Functional expression of the gene encoding cytidine triphosphate synthetase from Plasmodium falciparum which contains two novel sequences that are potential antimalarial targets. Mol Biochem Parasitol. 2005;143(2):200-8doi: 10.1016/j.molbiopara.2005.06.004.
Yuan, P., Hendriks, E. F., Fernandez, H. R., O'Sullivan, W. J., & Stewart, T. S. (2005). Functional expression of the gene encoding cytidine triphosphate synthetase from Plasmodium falciparum which contains two novel sequences that are potential antimalarial targets. Molecular and biochemical parasitology, 143(2), 200-8. https://doi.org/10.1016/j.molbiopara.2005.06.004
Yuan, Ping, et al. "Functional expression of the gene encoding cytidine triphosphate synthetase from Plasmodium falciparum which contains two novel sequences that are potential antimalarial targets." Molecular and biochemical parasitology vol. 143,2 (2005): 200-8. doi: https://doi.org/10.1016/j.molbiopara.2005.06.004
Yuan P, Hendriks EF, Fernandez HR, O'Sullivan WJ, Stewart TS. Functional expression of the gene encoding cytidine triphosphate synthetase from Plasmodium falciparum which contains two novel sequences that are potential antimalarial targets. Mol Biochem Parasitol. 2005 Oct;143(2):200-8. doi: 10.1016/j.molbiopara.2005.06.004. PMID: 16051382.
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Mol Biochem Parasitol. 2005 Oct;143(2):200-8. doi: 10.1016/j.molbiopara.2005.06.004.

Functional expression of the gene encoding cytidine triphosphate synthetase from Plasmodium falciparum which contains two novel sequences that are potential antimalarial targets.

Molecular and biochemical parasitology

Ping Yuan, Edward F Hendriks, Harvey R Fernandez, William J O'Sullivan, Thomas S Stewart

Affiliations

  1. School of Biotechnology and Biomolecular Sciences, University of New South Wales, Sydney, NSW 2052, Australia. [email protected]

PMID: 16051382 DOI: 10.1016/j.molbiopara.2005.06.004

Abstract

CTP synthetase (E C 6.3.4.2 UTP: ammonia ligase (ADP-forming)) catalyses the formation of CTP from UTP and, in the human parasite Plasmodium falciparum, is the sole source of cytidine nucleotides. It is thus a potential chemotherapeutic target, especially as the gene sequence indicated that the encoded GAT-domain of the enzyme contains two extended peptide segments (42aa and 223aa as compared to the host enzyme). Here, we circumvent the codon usage problems associated with the high A/T content of the P. falciparum sequence, especially evident in sequences encoding the extra peptides, to successfully express active recombinant P. falciparum CTP synthetase using preferred E. coli codons. This partially synthetic gene produced recombinant enzyme, containing the additional segments, which was functionally assayed for activity in vitro. We also show the native enzyme contains the additional peptides using immunoblots with antibodies derived from the recombinant protein. Confocal microscopy, using antibodies to the recombinant protein, provided evidence that the enzyme is expressed in vivo. This establishes for the first time that P. falciparum contain active CTP synthetase and that this enzyme contains two novel insert sequences in the functional enzyme.

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