An atypical mitogen-activated protein kinase (MAPK) homologue expressed ingametocytes of the human malaria parasite Plasmodium falciparum - Identification of a MAPK signature

The cDNA encoding Pfmap-2, an enzyme of the human malaria parasite Plasmodi um falciparum, was cloned, sequenced, and expressed in Escherichia coli, Th e open reading frame carried by the Pfmap-2 cDNA encodes a 508-amino acid p olypeptide of 59.2 kDa with maximal homology to mitogen-activated protein k inases (MAPKs) from various organisms. The purified recombinant enzyme disp layed functional characteristics of MAPKs such as (i) ability to undergo au tophosphorylation, (ii) ability to phosphorylate myelin basic protein, a cl assical MAPK substrate, (iii) regulation of kinase activity by a MAPK-speci fic phosphatase, and (iv) ability to be activated by component(s) present i n cell extracts. Mutational analysis of the recombinant protein allowed the identification of residues that are important for enzymatic activity. Nort hern blot analysis and immunofluorescence assays indicated that Pfmap-2 is expressed specifically in gametocytes, the form that is responsible for tra nsmission of the parasite to the mosquito vector. Gametocyte extracts activ ated recombinant Pfmap-2 more efficiently than extracts from asexual parasi tes, which is consistent with this stage specificity, Despite its overall h igh level of homology to MAPKs, Pfmap-2 presents the peculiarity of not pos sessing the conserved threonine-X-tyrosine activation motif usually found i n enzymes of this family; instead, it has a threonine-serine-histidine at t he same location. This atypical feature formed the basis for a detailed ana lysis of the primary structure of MAPKs, allowing us to define an operation al MAPK signature, which is shared by Pfmap-2. The fact that no MAPK from v ertebrates diverge in the activation motif suggests that the fine mechanism s of Pfmap-2 regulation may offer an opportunity for antimalarial drug targ eting.