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Characterization of a non‐mitochondrial type I phosphatidylserine decarboxylase in Plasmodium falciparum
Author(s) -
Baunaure Françoise,
Eldin Patrick,
Cathiard AnneMarie,
Vial Henri
Publication year - 2004
Publication title -
molecular microbiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.857
H-Index - 247
eISSN - 1365-2958
pISSN - 0950-382X
DOI - 10.1046/j.1365-2958.2003.03822.x
Subject(s) - biology , complementary dna , microbiology and biotechnology , endoplasmic reticulum , biochemistry , phosphatidylserine , peptide sequence , phosphatidylethanolamine , plasmodium falciparum , cdna library , gene , phospholipid , membrane , phosphatidylcholine , malaria , immunology
Summary In 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 µM and a V MAX of 680 ± 49 nmol of phosphatidylethanolamine formed h −1 mg −1 protein. Site‐directed mutagenesis of the VGSS active site demonstrated that the PfPSD proenzyme was processed into two non‐identical subunits (α and β) 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.