Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0024530 (malaria)
 44,886 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The plasmepsins are involved in the degradation of host cell hemoglobin during malaria infection. Plasmepsin I (PM I) initiates the degradative process, and has been suggested as an attractive target for the treatment of malaria. The production of active recombinant PM I, however, has been challenging. We report for the first time, the expression and partial characterization of soluble recombinant PM I from Plasmodium falciparum in which a truncated form of PM I (Lys77P-Leu329) (P indicates a propart residues) was fused to thioredoxin in the pET32b(+) vector, Trx-tPM I and expressed in Escherichia coli Rosetta-gami B (DE3)pLysS. The soluble fusion protein was purified from cell culture using a combination of Ni(2+) affinity and gel filtration chromatography and was capable of autocatalytic activation at pH 4.0-5.5, which occurred at Leu116P-Ser117P, seven residues upstream of the native cleavage site (Gly123P-Asn1). The mature tPM I (mtPM I) was capable of hydrolyzing both human hemoglobin with a pH optimum of pH 2.8-4.0 and the synthetic fluorogenic peptide EDANS-CO-CH(2)-CH(2)-CO-ALERMFLSFP-Dap(DABCYL)-OH with a dual pH optima of pH 2.5-3.0 and pH 4.5-5.5. Using the synthetic substrate, mtPM I exhibited kinetic parameters comparable to native PM I.
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PMID:Expression and enzymatic characterization of the soluble recombinant plasmepsin I from Plasmodium falciparum. 1807 24

Malaria kills >1 million people each year, in particular in sub-Saharan Africa. Although asexual forms are directly responsible for disease and death, sexual stages account for the transmission of Plasmodium parasites from human to the mosquito vector and therefore the spread of the parasite in the population. Development of a malaria vaccine is urgently needed to reduce morbidity and mortality. Vaccines against sexual stages of Plasmodium falciparum are meant to decrease the force of transmission and consequently reduce malaria burden. Pfs48/45 is specifically expressed in sexual stages and is a well established transmission-blocking (TB) vaccine candidate. However, production of correctly folded recombinant Pfs48/45 protein with display of its TB epitopes has been a major challenge. Here, we show the production of a properly folded Pfs48/45 C-terminal fragment by simultaneous coexpression with four periplasmic folding catalysts in Escherichia coli. This C-terminal fragment fused to maltose binding protein was produced at medium scale with >90% purity and a stability over at least a 9-month period. It induces uniform and high antibody titers in mice and elicits functional TB antibodies in standard membrane feeding assays in 90% of the immunized mice. Our data provide a clear perspective on the clinical development of a Pfs48/45-based TB malaria vaccine.
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PMID:Correctly folded Pfs48/45 protein of Plasmodium falciparum elicits malaria transmission-blocking immunity in mice. 1833 22

Antibodies from malaria-exposed individuals can agglutinate merozoites released from Plasmodium schizonts, thereby preventing them from invading new erythrocytes. Merozoite coat proteins attached to the plasma membrane are major targets for host antibodies and are therefore considered important malaria vaccine candidates. Prominent among these is the abundant glycosylphosphatidylinositol (GPI)-anchored merozoite surface protein 1 (MSP1) and particularly its C-terminal fragment (MSP1(19)) comprised of two epidermal growth factor (EGF)-like modules. In this paper, we revisit the role of agglutination and immunity using transgenic fluorescent marker proteins. We describe expression of heterologous MSP1(19)'miniproteins' on the surface of Plasmodium falciparum merozoites. To correctly express these proteins, we determined that GPI-anchoring and the presence of a signal sequence do not allow default export of proteins from the endoplasmic reticulum to merozoite surface and that extra sequence elements are required. The EGFs are insufficient for correct trafficking unless they are fused to additional residues that normally reside upstream of this fragment. Antibodies specifically targeting the surface-expressed miniprotein can inhibit erythrocyte invasion in vitro despite the presence of endogenous MSP1. Using a line expressing a green fluorescent protein-MSP1 fusion protein, we demonstrate that one mode of inhibition by antibodies targeting the MSP1(19) domain is the rapid agglutinating of merozoites prior to erythrocyte attachment.
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PMID:MSP1(19) miniproteins can serve as targets for invasion inhibitory antibodies in Plasmodium falciparum provided they contain the correct domains for cell surface trafficking. 1833 85

Falcipain-2 (FP-2), a papain family cysteine protease of Plasmodium falciparum, is a promising target for antimalarial chemotherapy. Designing inhibitors that are highly selective for falcipain-2 has been difficult because of broad specificity of different cysteine proteinases. Because propeptide regions of cysteine proteases have been shown to inhibit their cognate enzymes specifically and selectively, in the present study, we evaluated the inhibitory potential of few falcipain-2 proregion peptides. A 15 residue peptide (PP1) inhibited falcipain-2 enzyme activity in vitro. Studies on the uptake of PP1 into the parasitized erythrocytes showed access of peptide into the infected RBCs. PP1 fused with Antennapedia homeoprotein internalization domain blocked hemoglobin hydrolysis, merozoite release and markedly inhibited Plasmodium falciparum growth and maturation. Together, our results identify a peptide derived from the proregion of falcipain-2 that blocks late-stage malaria parasite development in RBCs, suggesting the development of peptide and peptidometric drugs against the human malaria parasite.
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PMID:A prodomain peptide of Plasmodium falciparum cysteine protease (falcipain-2) inhibits malaria parasite development. 1846 22

In red blood cells (RBCs) infected with the malaria parasite Plasmodium falciparum, a 19-residue region of the mature parasite-infected erythrocyte surface antigen (MESA) associates with RBC cytoskeleton protein 4.1R; an interaction essential for parasite survival. This region in MESA is adjacent to a host targeting motif found in other malaria parasite proteins exported to the membrane skeleton. To demonstrate function of these motifs in vivo, regions of MESA fused to a reporter were expressed in malaria parasites. Immunochemical analyses confirmed the requirement for both motifs in the trafficking and interaction of MESA with the cytoskeleton and demonstrates their function in vivo.
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PMID:In vivo studies support the role of trafficking and cytoskeletal-binding motifs in the interaction of MESA with the membrane skeleton of Plasmodium falciparum-infected red blood cells. 1848 75

Immunogenicity testing of Plasmodium falciparum antigens being considered as malaria vaccine candidates was undertaken in rabbits. The antigens compared were recombinant baculovirus MSP-1(19) and five Pichia pastoris candidates, including two versions of MSP-1(19), AMA-1 (domains I and II), AMA-1+MSP-1(19), and fused AMA-1/MSP-1(19)). Animals were immunized with equimolar amounts of each antigen, formulated in Montanide ISA720. The specificities and titers of antibodies were compared using immunofluorescence assays and enzyme-linked immunosorbent assay (ELISA). The antiparasite activity of immunoglobulin G (IgG) in in vitro cultures was determined by growth inhibition assay, flow cytometry, lactate dehydrogenase assay, and microscopy. Baculovirus MSP-1(19) immunizations produced the highest parasite-specific antibody titers in immunofluorescence assays. In ELISAs, baculovirus-produced MSP-1(19) induced more antibodies than any other single MSP-1(19) immunogen and three times more MSP-1(19) specific antibodies than the AMA-1/MSP-1(19) fusion. Antibodies induced by baculovirus MSP-1(19) gave the highest levels of growth inhibition in HB3 and 3D7 parasite cultures, followed by AMA-1+MSP-1(19) and the AMA-1/MSP-1(19) fusion. With the FCR3 isolate (homologous to the AMA-1 construct), antibodies to the three AMA-1-containing candidates gave the highest levels of growth inhibition at high IgG concentrations, but antibodies to baculovirus MSP-1(19) inhibited as well or better at lower IgG concentrations. The two P. pastoris-produced MSP-1(19)-induced IgGs conferred the lowest growth inhibition. Comparative analysis of immunogenicity of vaccine antigens can be used to prioritize candidates before moving to expensive GMP production and clinical testing. The assays used have given discriminating readouts but it is not known whether any of them accurately reflect clinical protection.
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PMID:Comparative testing of six antigen-based malaria vaccine candidates directed toward merozoite-stage Plasmodium falciparum. 1855 Jul 31

This study initially sought to investigate the immunostimulatory properties of recombinant adeno-associated virus (rAAV) with a view to developing a genetic vaccine for malaria using muscle as a target tissue. To augment humoral immunity, the AAV-encoded antigen was genetically fused with CTLA4-Ig, a recombinant molecule that binds B7 costimulatory molecules. At 10(9) vg, CTLA4-Ig fusion promoted the humoral immune response 100-fold and was dependent on CTLA4-Ig binding with B7 costimulatory molecules, confirming plasmid DNA models using this strategy. In distinct contrast, 10(12)-10(13) vg of rAAV1 specifically induced long-lived humoral tolerance through a mechanism that is independent of CTLA4-Ig binding with B7. This finding was unexpected, as rAAV delivery to muscle, unlike liver, has shown that this tissue provides a highly immunogenic environment for induction of humoral immunity against rAAV transgene products. An additional unpredicted consequence of antigen fusion with CTLA4-Ig was the enhancement of antigen expression by approximately one log, an effect mapped to the hinge and Fc domain of IgG(1,) but not involving antigen dimerization or the neonatal Fc receptor. Collectively, these findings significantly advance the potential of rAAV both as a vaccine or immunotherapeutic platform for the induction of antigen-specific humoral immunity or tolerance and as a gene therapeutic delivery system.
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PMID:Antigen-specific humoral tolerance or immune augmentation induced by intramuscular delivery of adeno-associated viruses encoding CTLA4-Ig-antigen fusion molecules. 1903 42

RTS,S is a pre-erythrocytic malaria vaccine candidate antigen based on the circumsporozoite surface protein of Plasmodium falciparum fused to HBsAg, incorporating a novel Adjuvant System (AS02). The first field efficacy of RTS,S/AS02 against infection was demonstrated in a trial initiated in The Gambia in 1998. This paper presents the five-year safety and immunogenicity follow up of the 306 men who were enrolled in the original trial. In the primary study men aged 18 to 45 years were randomized to receive either RTS,S/AS02 or rabies vaccine at 0, 1, 5 months followed by a booster dose at month 19. The subjects were observed for long term safety and immunogenicity continuously until month 58. Of the 153 subjects in each group at enrollment, 80 (52%) subjects in the RTS,S/AS02 group and 83 (54%) subjects in the rabies group returned for the final long-term follow-up visit at month 58. The main reason for non-attendance at month 58 was migration (76% of all drop-outs). Nine subjects in the RTS,S/AS02 group and seven in the rabies group experienced serious adverse events (SAEs) over the 58 month surveillance period, of which seven had a fatal outcome (five RTS,S/AS02 and two rabies group). None of the SAEs with fatal outcome were attributed to the study vaccine. Anti-CS antibody persistence compared to control was observed for five years, although titres had waned from post-booster levels; similar responses in anti-HBs antibody persistence were observed in initially HBsAg seronegative subjects. This study provides the first indication of the long-term safety and persistence of anti-CS and anti-HBs antibodies of the RTS,S vaccine candidate in combination with the novel AS02 Adjuvant System.
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PMID:Five-year safety and immunogenicity of GlaxoSmithKline's candidate malaria vaccine RTS,S/AS02 following administration to semi-immune adult men living in a malaria-endemic region of The Gambia. 1927 46

Malaria parasite-infected erythrocytes exhibit enhanced glucose utilisation and 6-phospho-1-fructokinase (PFK) is a key enzyme in glycolysis. Here we present the characterisation of PFK from the human malaria parasite Plasmodium falciparum. Of the two putative PFK genes on chromosome 9 (PfPFK9) and 11 (PfPFK11), only the PfPFK9 gene appeared to possess all the catalytic features appropriate for PFK activity. The deduced PfPFK proteins contain domains homologous to the plant-like pyrophosphate (PPi)-dependent PFK beta and alpha subunits, which are quite different from the human erythrocyte PFK protein. The PfPFK9 gene beta and alpha regions were cloned and expressed as His(6)- and GST-tagged proteins in Escherichia coli. Complementation of PFK-deficient E. coli and activity analysis of purified recombinant proteins confirmed that PfPFK9beta possessed catalytic activity. Monoclonal antibodies against the recombinant beta protein confirmed that the PfPFK9 protein has beta and alpha domains fused into a 200 kDa protein, as opposed to the independent subunits found in plants. Despite an overall structural similarity to plant PPi-PFKs, the recombinant protein and the parasite extract exhibited only ATP-dependent enzyme activity, and none with PPi. Unlike host PFK, the Plasmodium PFK was insensitive to fructose-2,6-bisphosphate (F-2,6-bP), phosphoenolpyruvate (PEP) and citrate. A comparison of the deduced PFK proteins from several protozoan PFK genome databases implicates a unique class of ATP-dependent PFK present amongst the apicomplexan protozoans.
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PMID:Plant-like phosphofructokinase from Plasmodium falciparum belongs to a novel class of ATP-dependent enzymes. 1950 69

Cholera and malaria are major diseases causing high mortality. The only licensed cholera vaccine is expensive; immunity is lost in children within 3 years and adults are not fully protected. No vaccine is yet available for malaria. Therefore, in this study, the cholera toxin-B subunit (CTB) of Vibrio cholerae fused to malarial vaccine antigens apical membrane antigen-1 (AMA1) and merozoite surface protein-1 (MSP1) was expressed in lettuce and tobacco chloroplasts. Southern blot analysis confirmed homoplasmy and stable integration of transgenes. CTB-AMA1 and CTB-MSP1 fusion proteins accumulated up to 13.17% and 10.11% (total soluble protein, TSP) in tobacco and up to 7.3% and 6.1% (TSP) in lettuce, respectively. Nine groups of mice (n = 10/group) were immunized subcutaneously (SQV) or orally (ORV) with purified antigens or transplastomic tobacco leaves. Significant levels of antigen-specific antibody titres of immunized mice completely inhibited proliferation of the malarial parasite and cross-reacted with the native parasite proteins in immunoblots and immunofluorescence studies. Protection against cholera toxin challenge in both ORV (100%) and SQV (89%) mice correlated with CTB-specific titres of intestinal, serum IgA and IgG1 in ORV and only IgG1 in SQV mice, but no other immunoglobulin. Increasing numbers of interleukin-10(+) T cell but not Foxp3(+) regulatory T cells, suppression of interferon-gamma and absence of interleukin-17 were observed in protected mice, suggesting that immunity is conferred via the Tr1/Th2 immune response. Dual immunity against two major infectious diseases provided by chloroplast-derived vaccine antigens for long-term (>300 days, 50% of mouse life span) offers a realistic platform for low cost vaccines and insight into mucosal and systemic immunity.
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PMID:Chloroplast-derived vaccine antigens confer dual immunity against cholera and malaria by oral or injectable delivery. 2005 Oct 36


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