The Role of Hsp70s in the Development and Pathogenicity of Plasmodium falciparum.

The main agent of human malaria, the protozoa, Plasmodium falciparum is known to infect liver cells, subsequently invading the host erythrocyte, leading to the manifestation of clinical outcomes of the disease. As part of its survival in the human host, P. falciparum employs several heat shock protein (Hsp) families whose primary purpose is to ensure cytoprotection through their molecular chaperone role. The parasite expresses six Hsp70s that localise to various subcellular organelles of the parasite, with one, PfHsp70-x, being exported to the infected human erythrocyte. The role of these Hsp70s in the survival and pathogenicity of malaria has received immense research attention. Several studies have reported on their structure-function features, network partnerships, and elucidation of their potential substrates. Apart from their role in cytoprotection and pathogenicity, Hsp70s are implicated in antimalarial drug resistance. As such, they are deemed potential antimalarial drug candidates, especially suited for co-targeting in combination therapies. In addition, Hsp70 is implicated in host immune modulation. The current report highlights the various structure-function features of these proteins, their roles in the development of malaria, current and prospective efforts being employed towards targeting them in malaria intervention efforts.

Heat Shock Proteins of Malaria: Highlights and Future Prospects.

The deadliest malaria parasite of humans, Plasmodium falciparum, is an obligate parasite that has had to develop mechanisms for survival under the unfavourable conditions it confronts within host cells. The chapters in the book "Heat Shock Proteins of Malaria" provide a critique of the evidence that heat shock proteins (Hsps) play a key role in the survival of P. falciparum in host cells. The role of the plasmodial Hsp arsenal is not limited to the protection of the parasite cell (largely through their role as molecular chaperones), as some of these proteins also promote the pathological development of malaria. This is largely due to the export of a large number of these proteins into the infected erythrocyte cytosol. Although P. falciparum erythrocyte membrane protein 1 (PfEMP1) is the main virulence factor for the malaria parasite, some of the exported plasmodial Hsps appear to augment parasite virulence. While this book largely delves into experimentally validated information on the role of Hsps in the development and pathogenicity of malaria, some of the information is based on hypotheses yet to be fully tested. Therefore, here we highlight what we know to be definite roles of plasmodial Hsps. Furthermore, we distill information that could provide practical insights on the options available for future research directions, including interventions against malaria that may target the role of Hsps in the development of the disease.

Bioprospecting for Novel Heat Shock Protein Modulators: The New Frontier for Antimalarial Drug Discovery?

Heat shock proteins are conserved molecules whose main role is to facilitate protein folding. However, they are also implicated in protein trafficking, protein assembly/disassembly, and functional maturation of proteins implicated in several biochemical pathways, including signal transduction. The role of heat shock proteins in the development of malaria parasites has recently become a subject of enormous interest. This is they do not only serve a cytoprotective role to ensure parasite survival but are implicated in the trafficking of several parasite proteins that are exported to the infected host red blood cell. Indeed, several heat shock proteins are also exported to the infected human red blood cell. In light of this, heat shock proteins along with other molecules are thought to modify the host cell, thus regulating the pathogenicity of malaria parasites. Even more important is their role in augmenting parasite resistance against antimalarial drugs. In light of the essential functions of several of these molecules in the development of malaria parasites, coupled with their role in antimalarial drug resistance, there is growing interest to target them as part of antimalarial drug discovery efforts. Several antimalarial compounds used so far originate from natural products. It is only logical that in our pursuit to identify small molecule inhibitors targeting heat shock proteins of malaria parasites, we turn to nature for answers and possible clues. In the current narrative, we focus attention on features of heat shock proteins of malaria parasites that make them amenable to targeting. In addition, we discuss various plant products that have been identified as sources of antimalarial compounds that target heat shock proteins. The current narrative seeks to inspire novel drug discovery experts, especially those working on natural compounds to focus on heat shock proteins as possible antimalarial targets. We further discuss the challenges of taking this route as part of our growing arsenal against malaria.

Characterization of an Atypical Trypanosoma brucei Hsp70 Demonstrates Its Cytosolic-Nuclear Localization and Modulation by Quercetin and Methylene Blue.

Trypanosoma brucei (Tb) harbours twelve Hsp70 chaperones. Of these, four are predicted to reside in the parasite cytosol. TbHsp70.c is predicted to be cytosolic and upregulated upon heat stress and is an ATPase that exhibits holdase chaperone function. Cytosol-localized Tbj2 stimulates the ATPase activity of TbHsp70.c. In the current study, immunofluorescence confirmed that TbHsp70.c is both a cytosolic and a nuclear protein. Furthermore, in silico analysis was used to elucidate an atypical linker and hydrophobic pocket. Tellingly, TbHsp70.c lacks the EEVD and GGMP motifs, both of which are implicated in substrate selectivity and co-chaperone binding in canonical Hsp70s. Far western analysis revealed that TbSTi1 interacts directly with TbHsp70 and TbHsp70.4, but does not bind TbHsp70.c. We further investigated the effect of quercetin and methylene blue on the Tbj2-driven ATPase activity of TbHsp70.c. We established that quercetin inhibited, whilst methylene blue enhanced, the Tbj2-stimulated ATPase activity of TbHsp70.c. Furthermore, these inhibitors were lethal to parasites. Lastly, we used molecular docking to show that quercetin and methylene blue may bind the nucleotide binding pocket of TbHsp70.c. Our findings suggest that small molecule inhibitors that target TbHsp70.c could be developed to serve as possible drug candidates against T. brucei.

Mutation of GGMP Repeat Segments of Plasmodium falciparum Hsp70-1 Compromises Chaperone Function and Hop Co-Chaperone Binding.

Parasitic organisms especially those of the Apicomplexan phylum, harbour a cytosol localised canonical Hsp70 chaperone. One of the defining features of this protein is the presence of GGMP repeat residues sandwiched between α-helical lid and C-terminal EEVD motif. The role of the GGMP repeats of Hsp70s remains unknown. In the current study, we introduced GGMP mutations in the cytosol localised Hsp70-1 of Plasmodium falciparum (PfHsp70-1) and a chimeric protein (KPf), constituted by the ATPase domain of E. coli DnaK fused to the C-terminal substrate binding domain of PfHsp70-1. A complementation assay conducted using E. coli dnaK756 cells demonstrated that the GGMP motif was essential for chaperone function of the chimeric protein, KPf. Interestingly, insertion of GGMP motif of PfHsp70-1 into DnaK led to a lethal phenotype in E. coli dnaK756 cells exposed to elevated growth temperature. Using biochemical and biophysical assays, we established that the GGMP motif accounts for the elevated basal ATPase activity of PfHsp70-1. Furthermore, we demonstrated that this motif is important for interaction of the chaperone with peptide substrate and a co-chaperone, PfHop. Our findings suggest that the GGMP may account for both the specialised chaperone function and reportedly high catalytic efficiency of PfHsp70-1.

Proteomic analysis of Plasmodium falciparum histone deacetylase 1 complex proteins.

Plasmodium falciparum histone deacetylases (PfHDACs) are an important class of epigenetic regulators that alter protein lysine acetylation, contributing to regulation of gene expression and normal parasite growth and development. PfHDACs are therefore under investigation as drug targets for malaria. Despite this, our understanding of the biological roles of these enzymes is only just beginning to emerge. In higher eukaryotes, HDACs function as part of multi-protein complexes and act on both histone and non-histone substrates. Here, we present a proteomics analysis of PfHDAC1 immunoprecipitates, identifying 26 putative P. falciparum complex proteins in trophozoite-stage asexual intraerythrocytic parasites. The co-migration of two of these (P. falciparum heat shock proteins 70-1 and 90) with PfHDAC1 was validated using Blue Native PAGE combined with Western blot. These data provide a snapshot of possible PfHDAC1 interactions and a starting point for future studies focused on elucidating the broader function of PfHDACs in Plasmodium parasites.

Small Molecule Inhibitors Targeting the Heat Shock Protein System of Human Obligate Protozoan Parasites.

Obligate protozoan parasites of the kinetoplastids and apicomplexa infect human cells to complete their life cycles. Some of the members of these groups of parasites develop in at least two systems, the human host and the insect vector. Survival under the varied physiological conditions associated with the human host and in the arthropod vectors requires the parasites to modulate their metabolic complement in order to meet the prevailing conditions. One of the key features of these parasites essential for their survival and host infectivity is timely expression of various proteins. Even more importantly is the need to keep their proteome functional by maintaining its functional capabilities in the wake of physiological changes and host immune responses. For this reason, molecular chaperones (also called heat shock proteins)-whose role is to facilitate proteostasis-play an important role in the survival of these parasites. Heat shock protein 90 (Hsp90) and Hsp70 are prominent molecular chaperones that are generally induced in response to physiological stress. Both Hsp90 and Hsp70 members are functionally regulated by nucleotides. In addition, Hsp70 and Hsp90 cooperate to facilitate folding of some key proteins implicated in cellular development. In addition, Hsp90 and Hsp70 individually interact with other accessory proteins (co-chaperones) that regulate their functions. The dependency of these proteins on nucleotide for their chaperone function presents an Achille's heel, as inhibitors that mimic ATP are amongst potential therapeutic agents targeting their function in obligate intracellular human parasites. Most of the promising small molecule inhibitors of parasitic heat shock proteins are either antibiotics or anticancer agents, whose repurposing against parasitic infections holds prospects. Both cancer cells and obligate human parasites depend upon a robust protein quality control system to ensure their survival, and hence, both employ a competent heat shock machinery to this end. Furthermore, some inhibitors that target chaperone and co-chaperone networks also offer promising prospects as antiparasitic agents. The current review highlights the progress made so far in design and application of small molecule inhibitors against obligate intracellular human parasites of the kinetoplastida and apicomplexan kingdoms.

Structural and biochemical characterization of Plasmodium falciparum Hsp70-x reveals functional versatility of its C-terminal EEVN motif.

Plasmodium falciparum, the main agent of malaria expresses six members of the heat shock protein 70 (Hsp70) family. Hsp70s serve as protein folding facilitators in the cell. Amongst the six Hsp70 species that P. falciparum expresses, Hsp70-x (PfHsp70-x), is partially exported to the host red blood cell where it is implicated in host cell remodeling. Nearly 500 proteins of parasitic origin are exported to the parasite-infected red blood cell (RBC) along with PfHsp70-x. The role of PfHsp70-x in the infected human RBC remains largely unclear. One of the defining features of PfHsp70-x is the presence of EEVN residues at its C-terminus. In this regard, PfHsp70-x resembles canonical eukaryotic cytosol-localized Hsp70s which possess EEVD residues at their C-termini in place of the EEVN residues associated with PfHsp70-x. The EEVD residues of eukaryotic Hsp70s facilitate their interaction with co-chaperones. Characterization of the role of the EEVN residues of PfHsp70-x could provide insights into the function of this protein. In the current study, we expressed and purified recombinant PfHsp70-x (full length) and its EEVN minus form (PfHsp70-xT ). We then conducted structure- function assays towards establishing the role of the EEVN motif of PfHsp70-x. Our findings suggest that the EEVN residues of PfHsp70-x are important for its ATPase activity and chaperone function. Furthermore, the EEVN residues are crucial for the direct interaction between PfHsp70-x and human Hsp70-Hsp90 organizing protein (hHop) in vitro. Hop facilitates functional cooperation between Hsp70 and Hsp90. However, it remains to be established if PfHsp70-x and hHsp90 cooperate in vivo.

Heat Shock Proteins as Immunomodulants.

Heat shock proteins (Hsps) are conserved molecules whose main role is to facilitate folding of other proteins. Most Hsps are generally stress-inducible as they play a particularly important cytoprotective role in cells exposed to stressful conditions. Initially, Hsps were generally thought to occur intracellulary. However, recent work has shown that some Hsps are secreted to the cell exterior particularly in response to stress. For this reason, they are generally regarded as danger signaling biomarkers. In this way, they prompt the immune system to react to prevailing adverse cellular conditions. For example, their enhanced secretion by cancer cells facilitate targeting of these cells by natural killer cells. Notably, Hsps are implicated in both pro-inflammatory and anti-inflammatory responses. Their effects on immune cells depends on a number of aspects such as concentration of the respective Hsp species. In addition, various Hsp species exert unique effects on immune cells. Because of their conservation, Hsps are implicated in auto-immune diseases. Here we discuss the various metabolic pathways in which various Hsps manifest immune modulation. In addition, we discuss possible experimental variations that may account for contradictory reports on the immunomodulatory function of some Hsps.

(-)-Epigallocatechin-3-Gallate Inhibits the Chaperone Activity of Plasmodium falciparum Hsp70 Chaperones and Abrogates Their Association with Functional Partners.

Heat shock proteins (Hsps), amongst them, Hsp70 and Hsp90 families, serve mainly as facilitators of protein folding (molecular chaperones) of the cell. The Hsp70 family of proteins represents one of the most important molecular chaperones in the cell. Plasmodium falciparum, the main agent of malaria, expresses six Hsp70 isoforms. Two (PfHsp70-1 and PfHsp70-z) of these localize to the parasite cytosol. PHsp70-1 is known to occur in a functional complex with another chaperone, PfHsp90 via a co-chaperone, P. falciparum Hsp70-Hsp90 organising protein (PfHop). (-)-Epigallocatechin-3-gallate (EGCG) is a green tea constituent that is thought to possess antiplasmodial activity. However, the mechanism by which EGCG exhibits antiplasmodial activity is not fully understood. A previous study proposed that EGCG binds to the N-terminal ATPase domain of Hsp70. In the current study, we overexpressed and purified recombinant forms of two P. falciparum cytosol localized Hsp70s (PfHsp70-1 and PfHsp70-z), and PfHop, a co-chaperone of PfHsp70-1. Using the surface plasmon resonance approach, we demonstrated that EGCG directly binds to the two Hsp70s. We further observed that binding of EGCG to the two proteins resulted in secondary and tertiary conformational changes. In addition, EGCG inhibited the ATPase and chaperone function of the two proteins. Furthermore, EGCG abrogated association of the two Hsp70s with their functional partners. Using parasites cultured in vitro at the blood stages, we observed that 2.9 µM EGCG suppressed 50% P. falciparum parasite growth (IC50). Our findings demonstrate that EGCG directly binds to PfHsp70-1 and PfHsp70-z to inhibit both the ATPase and chaperone functions of the proteins. Our study constitutes the first direct evidence suggesting that the antiplasmodial activity of EGCG is at least in part accounted for by its inhibition of Hsp70 function.

Extracts Obtained from Pterocarpus angolensis DC and Ziziphus mucronata Exhibit Antiplasmodial Activity and Inhibit Heat Shock Protein 70 (Hsp70) Function.

Malaria parasites are increasingly becoming resistant to currently used antimalarial therapies, therefore there is an urgent need to expand the arsenal of alternative antimalarial drugs. In addition, it is also important to identify novel antimalarial drug targets. In the current study, extracts of two plants, Pterocarpus angolensis and Ziziphus mucronata were obtained and their antimalarial functions were investigated. Furthermore, we explored the capability of the extracts to inhibit Plasmodium falciparum heat shock protein 70 (Hsp70) function. Heat shock protein 70 (Hsp70) are molecular chaperones whose function is to facilitate protein folding. Plasmodium falciparum the main agent of malaria, expresses two cytosol-localized Hsp70s: PfHsp70-1 and PfHsp70-z. The PfHsp70-z has been reported to be essential for parasite survival, while inhibition of PfHsp70-1 function leads to parasite death. Hence both PfHsp70-1 and PfHsp70-z are potential antimalarial drug targets. Extracts of P. angolensis and Z. mucronata inhibited the basal ATPase and chaperone functions of the two parasite Hsp70s. Furthermore, fractions of P. angolensis and Z. mucronata inhibited P. falciparum 3D7 parasite growth in vitro. The extracts obtained in the current study exhibited antiplasmodial activity as they killed P. falciparum parasites maintained in vitro. In addition, the findings further suggest that some of the compounds in P. angolensis and Z. mucronata may target parasite Hsp70 function.

Swapping the linkers of canonical Hsp70 and Hsp110 chaperones compromises both self-association and client selection.

Plasmodium falciparum heat shock protein 70-1 (PfHsp70-1) and PfHsp70-z are essential cytosol localised chaperones of the malaria parasite. The two chaperones functionally interact to drive folding of several parasite proteins. While PfHsp70-1 is regarded as a canonical Hsp70 chaperone, PfHsp70-z belongs to the Hsp110 subcluster. One of the distinctive features of PfHsp70-z is its unique linker segment which delineates it from canonical Hsp70. In the current study, we elucidated the role of the linker in regulating Hsp70 self-association and client selection. Using recombinant forms of PfHsp70-1, PfHsp70-z and E. coli Hsp70 (DnaK) and their respective linker switch mutants we investigated self-association of the chaperones using surface plasmon resonance (SPR) analysis. The effect of the changes on client selectivity was investigated on DnaK and its mutant through co-affinity chromatography coupled to LC-MS analysis. Our findings demonstrated that the linker is important for both Hsp70 self-association and client binding.

Iso-mukaadial acetate and ursolic acid acetate bind to Plasmodium Falciparum heat shock protein 70: towards targeting parasite protein folding pathway.

Plasmodium falciparum is the most lethal malaria parasite. P. falciparum Hsp70 (PfHsp70) is an essential molecular chaperone (facilitates protein folding) and is deemed a prospective antimalarial drug target. The present study investigates the binding capabilities of select plant derivatives, iso-mukaadial acetate (IMA) and ursolic acid acetate (UAA), against P. falciparum using an in silico docking approach. The interaction between the ligands and PfHsp70 was evaluated using plasmon resonance (SPR) analysis. Molecular docking, binding free energy analysis and molecular dynamics simulations were conducted towards understanding the mechanisms by which the compounds bind to PfHsp70. The molecular docking results revealed ligand flexibilities, conformations and positions of key amino acid residues and protein-ligand interactions as crucial factors accounting for selective inhibition of Hsp70. The simulation results also suggest protein-ligand van der Waals forces as the driving force guiding the interaction of these compounds with PfHsp70. Of the two compounds, UAA and IMA bound to PfHsp70 within the micromolar range based on surface plasmon resonance (SPR) based binding assay. Our findings pave way for future rational design of new selective compounds targeting PfHsp70.

Escherichia coli-based Complementation Assay to Study the Chaperone Function of Heat Shock Protein 70.

Heat shock protein 70 (Hsp70) is a conserved protein that facilitates the folding of other proteins within the cell, making it a molecular chaperone. While Hsp70 is not essential for E. coli cells growing under normal conditions, this chaperone becomes indispensable for growth at elevated temperatures. Since Hsp70 is highly conserved, one way to study the chaperone function of Hsp70 genes from various species is to heterologously express them in E. coli strains that are either deficient in Hsp70 or express a native Hsp70 that is functionally compromised. E. coli dnaK756 cells are unable to support λ bacteriophage DNA. Furthermore, their native Hsp70 (DnaK) exhibits elevated ATPase activity while demonstrating reduced affinity for GrpE (Hsp70 nucleotide exchange factor). As a result, E. coli dnaK756 cells grow adequately at temperatures ranging from 30 °C to 37 °C, but they die at elevated temperatures (>40 °C). For this reason, these cells serve as a model for studying the chaperone activity of Hsp70. Here, we describe a detailed protocol for the application of these cells to conduct a complementation assay, enabling the study of the in cellulo chaperone function of Hsp70.

Insertion of GGMP repeat residues of Plasmodium falciparum Hsp70-1 in the lid of DnaK adversely impacts client recognition.

Although Hsp70 is a conserved molecular chaperone, it exhibits some degree of functional specialisation across species. Features of Hsp70 regulating its functional specialisation remain to be fully established. We previously demonstrated that E. coli Hsp70 (DnaK) exhibits functional features that distinguishes it from PfHsp70-1, a canonical cytosolic Hsp70 of Plasmodium falciparum. One of the defining features of PfHsp70-1 is that it possesses GGMP repeat residues located in its C-terminal lid segment, while DnaK lacks this motif. Previously, we demonstrated that the insertion of GGMP repeat residues of PfHsp70-1 into E. coli DnaK abrogates the chaperone activity of DnaK. However, the role of the GGMP motif in regulating Hsp70 function remains to be fully understood. To explore the function of this motif, we expressed recombinant forms of wild type DnaK and its GGMP insertion motif, DnaK-G and systematically characterised the structure-function features of the two proteins using in silico analysis, biophysical approaches and an in cellulo complementation assay. Our findings demonstrated that the GGMP inserted in DnaK compromised various functional features such as nucleotide binding, allostery, substrate binding affinity and cellular proteome client selectivity. These findings thus, highlight the GGMP motif of Hsp70 as an important functional module.

Correction: Bopape et al. The Genome of a Pigeonpea Compatible Rhizobial Strain '10ap3' Appears to Lack Common Nodulation Genes. Genes 2023, 14, 1084.

Prof. Dr. Ahmed Idris Hassen was not included as an author in the original publication [...].

Ursolic acid acetate and iso-mukaadial acetate bind to Plasmodium falciparum Hsp90, abrogating its chaperone function in vitro.

Plasmodium falciparum is the most lethal malaria parasite. Increasing incidences of drug resistance of P. falciparum have prompted the need for discovering new and effective antimalarial compounds with an alternative mode of action. Heat shock protein 90 (PfHsp90) facilitates protein folding and is a promising antimalarial drug target. We have previously reported that iso-mukaadial acetate (IMA) and ursolic acid acetate (UAA) exhibit antimalarial activity. We investigated the abilities of IMA and UAA to bind PfHsp90 by molecular docking and dynamics simulations. The in silico predictions were validated by biochemical assays conducted on recombinant PfHsp90. The interaction between the ligands and PfHsp90 was evaluated using ultraviolet-visible spectroscopy (UV-vis), Fourier transform infrared (FTIR), and surface plasmon resonance (SPR) analysis. The results obtained by docking calculations and MD dynamics simulation predicted that UAA and IMA preferentially bound to PfHsp90 via the N-terminal domain, with UAA binding more stable than IMA. UV-vis-based data suggest that PfHsp90 harbors buried aromatic amino acids, which were exposed in the presence of either IMA or UAA. In addition, data obtained using FTIR suggested that IMA and UAA destabilized the secondary structure of PfHsp90. Of the two compounds, UAA bound to PfHsp90 within the micromolar range based on surface plasmon resonance (SPR)-based binding assay. Furthermore, both compounds disrupted the holdase chaperone function of PfHsp90 as the chaperone failed to suppress heat-induced aggregation of the model proteins, malate dehydrogenase (MDH), luciferase, and citrate synthase in vitro. In addition, both compounds lowered the ATPase activity of PfHsp90. The molecular dynamics simulation analysis indicated that the docked complexes were mostly stable for 100 ns, validating the data obtained through the biochemical assays. Altogether, this study expands the repository of antiplasmodial compounds that have PfHsp90 among their possible targets.

Plasmodium falciparum heat shock proteins as antimalarial drug targets: An update.

Global efforts to eradicate malaria are threatened by multiple factors, particularly the emergence of antimalarial drug resistant strains of Plasmodium falciparum. Heat shock proteins (HSPs), particularly P. falciparum HSPs (PfHSPs), represent promising drug targets due to their essential roles in parasite survival and virulence across the various life cycle stages. Despite structural similarities between human and malarial HSPs posing challenges, there is substantial evidence for subtle differences that could be exploited for selective drug targeting. This review provides an update on the potential of targeting various PfHSP families (particularly PfHSP40, PfHSP70, and PfHSP90) and their interactions within PfHSP complexes as a strategy to develop new antimalarial drugs. In addition, the need for a deeper understanding of the role of HSP complexes at the host-parasite interface is highlighted, especially heterologous partnerships between human and malarial HSPs, as this opens novel opportunities for targeting protein-protein interactions crucial for malaria parasite survival and pathogenesis.

Cysteine-capped gold nanoparticles suppress aggregation of proteins exposed to heat stress.

Gold nanoparticles show a lot of promise as potential agents for drug delivery and disease diagnosis. Because of this, it is important that the interaction between gold nanoparticles and biomolecules be well characterized to avoid undesirable consequences. In this study, gold nanoparticles were synthesized by the reduction of gold salt by sodium borohydride in the presence of cysteine as the capping agent. The physical features of the nanoparticles were analyzed using Ultraviolet-Visible spectrophotometry and transmission electron microscopy. The interaction between gold nanoparticles and the following proteins: bovine serum albumin, citrate synthase, malate dehydrogenase, and human heat shock protein 70 was investigated by UV-Vis spectrophotometry. The stability of the proteins against heat stress was assessed by monitoring their aggregation at 48 °C, either in the presence or absence of gold nanoparticles. The gold nanoparticles were capable of suppressing the heat-induced aggregation of the proteins. Furthermore, apart from possessing independent protein-aggregation suppression function, the AuNPs also augmented the chaperone function of human heat shock protein 70. Findings from this study demonstrate that cyteine-coated gold nanoparticles exhibit chaperone-like activity and have the capability to stabilize proteins to which they may be conjugated.

Anti-plasmodial activity of some Zulu medicinal plants and of some triterpenes isolated from them.

Mimusops caffra E. Mey. ex A.DC and Mimusops obtusifolia Lam (both members of the Sapotaceae family), and Hypoxis colchicifolia Bak (family Hypoxidaceae) are used by traditional healers in Zululand to manage malaria. Anti-plasmodial investigation of the crude extracts and some triterpenes isolated from the plants showed activity against a chloroquine sensitive (CQS) strain of Plasmodium falciparum (D10). Among the crude extracts the leaves of M. caffra exhibited the highest activity, with an IC50 of 2.14 µg/mL. The pentacyclic tritepenoid ursolic acid (1), isolated from the leaves of M. caffra was the most active compound (IC50 6.8 µg/mL) as compared to taraxerol (2) and sawamilletin (3) isolated from the stem bark of M. obtusifolia (IC50 > 100). Chemical modification of the ursolic acid (1) to 3ß-acetylursolic acid (4) greatly enhanced its anti-plasmodial activity. Compound 4 reduced parasitaemia against Plasmodium berghei by 94.01% in in vivo studies in mice. The cytotoxicity of 3ß-acetylursolic acid (IC50) to two human cell lines (HEK293 and HepG2) was 366.00 µg/mL and 566.09 µg/mL, respectively. The results validate the use of these plants in folk medicine.