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A fundamental problem in systems biology and whole genome sequence analysis is how to infer functions for the many uncharacterized proteins that are identified, whether they are conserved across organisms of different phyla or are phylum-specific. This problem is especially acute in pathogens, such as malaria parasites, where genetic and biochemical investigations are likely to be more difficult. Here we perform comparative expression analysis on Plasmodium parasite life cycle data derived from P. falciparum blood, sporozoite, zygote and ookinete stages, and P. yoelii mosquito oocyst and salivary gland sporozoites, blood and liver stages and show that type II fatty acid biosynthesis genes are upregulated in liver and insect stages relative to asexual blood stages. We also show that some universally uncharacterized genes with orthologs in Plasmodium species, Saccharomyces cerevisiae and humans show coordinated transcription patterns in large collections of human and yeast expression data and that the function of the uncharacterized genes can sometimes be predicted based on the expression patterns across these diverse organisms. We also use a comprehensive and unbiased literature mining method to predict which uncharacterized parasite-specific genes are likely to have roles in processes such as gliding motility, host-cell interactions, sporozoite stage, or rhoptry function. These analyses, together with proteinprotein interaction data, provide probabilistic models that predict the function of 926 uncharacterized malaria genes and also suggest that malaria parasites may provide a simple model system for the study of some human processes. These data also provide a foundation for further studies of transcriptional regulation in malaria parasites. [ABSTRACT FROM AUTHOR]

Summary: The interplay between ATP generating and utilizing pathways in a cell is responsible for maintaining cellular ATP/energy homeostasis that is reflected by Adenylate Energy Charge (AEC) ratio. Adenylate kinase (AK), that catalyzes inter‐conversion of ADP, ATP and AMP, plays a major role in maintaining AEC and is regulated by cellular AMP levels. Hence, the enzymes AMP deaminase (AMPD) and nucleotidases, which catabolize AMP, indirectly regulate AK activity and in‐turn affect AEC. Here, we present the first report on AMPD from Plasmodium, the causative agent of malaria. The recombinant enzyme expressed in Saccharomyces cerevisiae was studied using functional complementation assay and residues vital for enzyme activity have been identified. Similarities and differences between Plasmodium falciparum AMPD (PfAMPD) and its homologs from yeast, Arabidopsis and humans are also discussed. The AMPD gene was deleted in the murine malaria parasite P. berghei and was found to be dispensable during all stages of the parasite life cycle. However, when episomal expression was attempted, viable parasites were not obtained, suggesting that perturbing AMP homeostasis by over‐expressing AMPD might be lethal. As AMPD is known to be allosterically modulated by ATP, GTP and phosphate, allosteric activators of PfAMPD could be developed as anti‐parasitic agents. [ABSTRACT FROM AUTHOR]

Background: Emerging resistance of the malaria parasite Plasmodium to current therapies underscores the critical importance of exploring novel strategies for disease eradication. Plasmodium species are obligate intracellular protozoan parasites. They rely on an unusual form of substrate-dependent motility for their migration on and across host-cell membranes and for host cell invasion. This peculiar motility mechanism is driven by the 'glideosome', an actin-myosin associated, macromolecular complex anchored to the inner membrane complex of the parasite. Myosin A, actin, aldolase, and thrombospondin-related anonymous protein (TRAP) constitute the molecular core of the glideosome in the sporozoite, the mosquito stage that brings the infection into mammals. Methods: Virtual library screening of a large compound library against the PfAldolase-TRAP complex was used to identify candidate compounds that stabilize and prevent the disassembly of the glideosome. The mechanism of these compounds was confirmed by biochemical, biophysical and parasitological methods. Results: A novel inhibitory effect on the parasite was achieved by stabilizing a protein-protein interaction within the glideosome components. Compound 24 disrupts the gliding and invasive capabilities of Plasmodium parasites in in vitro parasite assays. A high-resolution, ternary X-ray crystal structure of PfAldolase-TRAP in complex with compound 24 confirms the mode of interaction and serves as a platform for future ligand optimization. Conclusion: This proof-of-concept study presents a novel approach to anti-malarial drug discovery and design. By strengthening a protein-protein interaction within the parasite, an avenue towards inhibiting a previously "undruggable" target is revealed and the motility motor responsible for successful invasion of host cells is rendered inactive. This study provides new insights into the malaria parasite cell invasion machinery and convincingly demonstrates that liver cell invasion is dramatically reduced by 95% in the presence of the small molecule stabilizer compound 24. [ABSTRACT FROM AUTHOR]

Background: Emerging resistance of the malaria parasite Plasmodium to current therapies underscores the critical importance of exploring novel strategies for disease eradication. Plasmodium species are obligate intracellular protozoan parasites. They rely on an unusual form of substrate-dependent motility for their migration on and across host-cell membranes and for host cell invasion. This peculiar motility mechanism is driven by the ‘glideosome’, an actin–myosin associated, macromolecular complex anchored to the inner membrane complex of the parasite. Myosin A, actin, aldolase, and thrombospondin-related anonymous protein (TRAP) constitute the molecular core of the glideosome in the sporozoite, the mosquito stage that brings the infection into mammals. Methods: Virtual library screening of a large compound library against the PfAldolase–TRAP complex was used to identify candidate compounds that stabilize and prevent the disassembly of the glideosome. The mechanism of these compounds was confirmed by biochemical, biophysical and parasitological methods. Results: A novel inhibitory effect on the parasite was achieved by stabilizing a protein–protein interaction within the glideosome components. Compound 24 disrupts the gliding and invasive capabilities of Plasmodium parasites in in vitro parasite assays. A high-resolution, ternary X-ray crystal structure of PfAldolase–TRAP in complex with compound 24 confirms the mode of interaction and serves as a platform for future ligand optimization. Conclusion: This proof-of-concept study presents a novel approach to anti-malarial drug discovery and design. By strengthening a protein–protein interaction within the parasite, an avenue towards inhibiting a previously “undruggable” target is revealed and the motility motor responsible for successful invasion of host cells is rendered inactive. This study provides new insights into the malaria parasite cell invasion machinery and convincingly demonstrates that liver cell invasion is dramatically reduced by 95 % in the presence of the small molecule stabilizer compound 24. [ABSTRACT FROM AUTHOR]