Identification and biochemical characterisation of a protein phosphatase 5 homologue from Plasmodium falciparum.
We report the identification of a new serine/threonine phosphatase from Plasmodium falciparum at the DNA and protein levels. A 1.8 kb cDNA fragment encoding the protein phosphatase was identified via PCR amplification. The sequence has a coding capacity of 594 amino acids. Immunoblot analysis of P. falciparum extracts showed that antibodies generated against the His(6)-fusion protein recognise a protein of approximately 80 kDa. The deduced amino acid sequence shares 55% identity with a mouse protein, identified as Protein Phosphatase 5 (PP5). We show that the P. falciparum PP5 homologue (PfPP5) has all structural and functional characteristics of this class of enzymes. It contains three tetratricopeptide repeats (TPR) and a nuclear targeting sequence at its N-terminus and a highly conserved C-terminal catalytic domain. Southern blot results are compatible with the existence of PfPP5 as a single copy gene. Purified recombinant protein, like the native protein enriched from P. falciparum extracts exhibited phosphatase activity that can be enhanced by both arachidonic and oleic acids, but not by myristic or stearic acid. In addition, the activity is inhibited by okadaic acid (OA) with an IC(50) of 4 nM. Immunofluorescence microscopy has localised PfPP5 preferentially to the nucleus. The function of PfPP5 is presently unclear, but like other PP5s of many eukaryotic organisms, it may have important regulatory functions in the parasite cell cycle.
Lindenthal
Christoph
C
Department of Parasitology, Institute for Tropical Medicine, University of Tübingen, Wilhelmstrasse 27, 72074 Tübingen, Germany.
Klinkert
Mo-Quen
MQ
D000595:Amino Acid Sequence; D000818:Animals; D016718:Arachidonic Acid / Q000494:pharmacology; D015153:Blotting, Western; D003001:Cloning, Molecular; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation / Q000187:drug effects; D018507:Gene Expression Regulation, Developmental; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D010749:Phosphoprotein Phosphatases / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D010963:Plasmodium falciparum / Q000201:enzymology*; D021381:Protein Transport; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017386:Sequence Homology, Amino Acid
D009687:Nuclear Proteins; D012333:RNA, Messenger; D011994:Recombinant Proteins; D016718:Arachidonic Acid; D010749:Phosphoprotein Phosphatases; C089641:protein phosphatase 5