Characterization of a non‐mitochondrial type I phosphatidylserine decarboxylase in Plasmodium falciparum

Abstract

SummaryIn search of key enzymes in Plasmodium phospholipid metabolism, we demonstrate the presence of a parasite-encoded phosphatidylserine decarboxylase (PSD) in the membrane fraction of Plasmodium falciparum -infected erythrocytes. PSD cDNA, encoding phosphatidylserine decarboxylase ( PfPSD ), was cloned by screening a directional cDNA library derived from the trophozoite erythrocytic stage. The corresponding PfPSD gene is located on chromosome 9 of P. falciparum , contains one intron of 938 nucleotides and is transcribed into a 3.7 kb mRNA. PfPSD cDNA encodes a putative protein of 362 amino acids, with a predicted molecular mass of 42.6 kDa, which clearly belongs to the type I PSD family. Only a 35 kDa polypeptide was detected in the parasite using a specific rabbit antiserum. PfPSD has a 314VGSS317 sequence near its carboxyl-terminus that is related to the Escherichia coli , yeast and human LGST motif, which is the site of proenzyme processing. PSD enzyme was expressed in E. coli with a K M of 63 ± ± ± ± 19 m m m m M and a V MAX of 680 ± ± ± ± 49 nmol of phosphatidylethanolamine formed h ----1 mg ----1 protein. Sitedirected mutagenesis of the VGSS active site demonstrated that the PfPSD proenzyme was processed into two non-identical subunits ( a a a a and b b b b ) and revealed the crucial role played by each residue in enzyme processing and activity. Using indirect immunofluorescence, PfPSD labelling was co-localized with an endoplasmic reticulum marker, but not with a mitochondrial vital dye. This P. falciparum PSD is the first type I PSD identified in the endoplasmic reticulum compartment.

Phosphatidylethanolamine Synthesis in the Parasite Mitochondrion Is Required for Efficient Growth but Dispensable for Survival of Toxoplasma gondii

“…Most type I PSD enzymes localize in mitochondria except for PfPSD1, which is reported to be expressed in the parasite ER (37). Unlike Plasmodium but consistent with mammalian, plant, and yeast cells (38 -40), we demonstrate a typically conserved mitochondrial expression of TgPSD1mt.…”

Identification of Gene Encoding Plasmodium knowlesi Phosphatidylserine Decarboxylase by Genetic Complementation in Yeast and Characterization of in Vitro Maturation of Encoded Enzyme

“…The sequence is not only required for the mitochondrial targeting but also for the correct processing into ␤ and ␣ fragments (33). Previously, plant PSD sequences from tomato and Arabidopsis were identified and expressed in the yeast psd1psd2 double mutant strain (10). The N-terminal mitochondrial targeting sequence of plant PSD protein was inhibitory to its functional expression in yeast.…”

“…P-Etn can also be methylated to form phosphocholine (P-Cho), which can be further metabolized to phosphatidylcholine (PtdCho) via a CDP-Cho intermediate. In the PtdSer decarboxylation pathway, serine is incorporated into phospholipid by a parasite PtdSer synthase and is subsequently decarboxylated to form PtdEtn by PtdSer decarboxylase (PfPSD) (10). Unlike yeast, Plasmodium lacks lipid methyltransferases for conversion of PtdEtn to PtdCho.…”

Comparative genomics and evolution of eukaryotic phospholipid biosynthesis

“…Disruption of both PSD1 and PSD2 genes causes yeast to become ethanolamine auxotrophs, whereas deletion of the single mouse PSD gene is embryonic lethal [140]. PSD genes have been also cloned and characterized from plants [141] and P. falciparum [142]. RNAi phenotypes associated with the single C. elegans PSD gene include abnormal embryonic and post-embryonic development, slow growth and an unusual transparent appearance of the worms [37,38,48,143].…”