2-Hydroxypropyl-ß-cyclodextrin (HPßCD) as a Potential Therapeutic Agent for Breast Cancer.

Cholesterol accumulation is documented in various malignancies including breast cancer. Consequently, depleting cholesterol in cancer cells can serve as a viable treatment strategy. We identified the potency of 2-hydroxypropyl-ß-cyclodextrin (HPßCD), a cholesterol-depletor in vitro against two breast cancer cell lines: MCF-7 (Oestrogen-receptor positive, ER+) and MDA-MB-231 (Triple negative breast cancer (TNBC)). The results were then compared against two non-cancerous cell lines using cytotoxic-, apoptosis-, and cholesterol-based assays. Treatment with HPßCD showed preferential and significant cytotoxic potential in cancer cells, inducing apoptosis in both cancer cell lines (p < 0.001). This was mediated due to significant depletion of cholesterol (p < 0.001). We further tested HPßCD in a MF-1 mice (n = 14) xenograft model and obtained 73.9%, 94% and 100% reduction in tumour size for late-, intermediate-, and early-stage TNBC, respectively. We also detected molecular-level perturbations in the expression patterns of several genes linked to breast cancer and cholesterol signalling pathways using RT2-PCR arrays and have identified SFRP1 as a direct binding partner to HPßCD through SPR drug interaction analysis. This work unravels mechanistic insights into HPßCD-induced cholesterol depletion, which leads to intrinsic apoptosis induction. Results from this study potentiate employing cholesterol depletion as a promising unconventional anticancer therapeutic strategy, which warrants future clinical investigations.

The Genome of a Pigeonpea Compatible Rhizobial Strain '10ap3' Appears to Lack Common Nodulation Genes.

The symbiotic fixation of atmospheric nitrogen (N) in root nodules of tropical legumes such as pigeonpea (Cajanus cajan) is a complex process, which is regulated by multiple genetic factors at the host plant genotype microsymbiont interface. The process involves multiple genes with various modes of action and is accomplished only when both organisms are compatible. Therefore, it is necessary to develop tools for the genetic manipulation of the host or bacterium towards improving N fixation. In this study, we sequenced the genome of a robust rhizobial strain, Rhizobium tropici '10ap3' that was compatible with pigeonpea, and we determined its genome size. The genome consisted of a large circular chromosome (6,297,373 bp) and contained 6013 genes of which 99.13% were coding sequences. However only 5833 of the genes were associated with proteins that could be assigned to specific functions. The genes for nitrogen, phosphorus and iron metabolism, stress response and the adenosine monophosphate nucleoside for purine conversion were present in the genome. However, the genome contained no common nod genes, suggesting that an alternative pathway involving a purine derivative was involved in the symbiotic association with pigeonpea.

Human granzyme B binds Plasmodium falciparum Hsp70-x and mediates antiplasmodial activity in vitro.

Cell surface-bound human Hsp70 (hHsp70) sensitises tumour cells to the cytolytic attack of natural killer (NK) cells through the mediation of apoptosis-inducing serine protease, granzyme B (GrB). hHsp70 is thought to recruit NK cells to the immunological synapse via the extracellularly exposed 14 amino acid sequence, TKDNNLLGRFELSG, known as the TKD motif of Hsp70. Plasmodium falciparum-infected red blood cells (RBCs) habour both hHsp70 and an exported parasite Hsp70 termed PfHsp70-x. Both PfHsp70-x and hHsp70 share conserved TKD motifs. The role of PfHsp70-x in facilitating GrB uptake in malaria parasite-infected RBCs remains unknown, but hHsp70 enables a perforin-independent uptake of GrB into tumour cells. In the current study, we comparatively investigated the direct binding of GrB to either PfHsp70-x or hHsp70 in vitro. Using ELISA, slot blot assay and surface plasmon resonance (SPR) analysis, we demonstrated a direct interaction of GrB with hHsp70 and PfHsp70-x. SPR analysis revealed a higher affinity of GrB for PfHsp70-x than hHsp70. In addition, we established that the TKD motif of PfHsp70-x directly interacts with GrB. The data further suggest that the C-terminal EEVN motif of PfHsp70-x augments the affinity of PfHsp70-x for GrB but is not a prerequisite for the binding. A potent antiplasmodial activity (IC50 of 0.5 µM) of GrB could be demonstrated. These findings suggest that the uptake of GrB by parasite-infected RBCs might be mediated by both hHsp70 and PfHsp70-x. The combined activity of both proteins could account for the antiplasmodial activity of GrB at the blood stage.

Inhibition of Plasmodium falciparum Hsp70-Hop partnership by 2-phenylthynesulfonamide.

Plasmodium falciparum Hsp70-1 (PfHsp70-1; PF3D7_0818900) and PfHsp90 (PF3D7_0708400) are essential cytosol localized chaperones of the malaria parasite. The two chaperones form a functional complex via the adaptor protein, Hsp90-Hsp70 organizing protein (PfHop [PF3D7_1434300]), which modulates the interaction of PfHsp70-1 and PfHsp90 through its tetracopeptide repeat (TPR) domains in a nucleotide-dependent fashion. On the other hand, PfHsp70-1 and PfHsp90 possess C-terminal EEVD and MEEVD motifs, respectively, which are crucial for their interaction with PfHop. By coordinating the cooperation of these two chaperones, PfHop plays an important role in the survival of the malaria parasite. 2-Phenylthynesulfonamide (PES) is a known anti-cancer agent whose mode of action is to inhibit Hsp70 function. In the current study, we explored the antiplasmodial activity of PES and investigated its capability to target the functions of PfHsp70-1 and its co-chaperone, PfHop. PES exhibited modest antiplasmodial activity (IC50 of 38.7 ± 0.7 µM). Furthermore, using surface plasmon resonance (SPR) analysis, we demonstrated that PES was capable of binding recombinant forms of both PfHsp70-1 and PfHop. Using limited proteolysis and intrinsic fluorescence-based analysis, we showed that PES induces conformational changes in PfHsp70-1 and PfHop. In addition, we demonstrated that PES inhibits the chaperone function of PfHsp70-1. Consequently, PES abrogated the association of the two proteins in vitro. Our study findings contribute to the growing efforts to expand the arsenal of potential antimalarial compounds in the wake of growing parasite resistance against currently used drugs.

Host cell stress response as a predictor of COVID-19 infectivity and disease progression.

The coronavirus disease (COVID-19) caused by a coronavirus identified in December 2019 has caused a global pandemic. COVID-19 was declared a pandemic in March 2020 and has led to more than 6.3 million deaths. The pandemic has disrupted world travel, economies, and lifestyles worldwide. Although vaccination has been an effective tool to reduce the severity and spread of the disease there is a need for more concerted approaches to fighting the disease. COVID-19 is characterised as a severe acute respiratory syndrome . The severity of the disease is associated with a battery of comorbidities such as cardiovascular diseases, cancer, chronic lung disease, and renal disease. These underlying diseases are associated with general cellular stress. Thus, COVID-19 exacerbates outcomes of the underlying conditions. Consequently, coronavirus infection and the various underlying conditions converge to present a combined strain on the cellular response. While the host response to the stress is primarily intended to be of benefit, the outcomes are occasionally unpredictable because the cellular stress response is a function of complex factors. This review discusses the role of the host stress response as a convergent point for COVID-19 and several non-communicable diseases. We further discuss the merits of targeting the host stress response to manage the clinical outcomes of COVID-19.

Characterisation of a unique linker segment of the Plasmodium falciparum cytosol localised Hsp110 chaperone.

Plasmodium falciparum expresses two essential cytosol localised chaperones; PfHsp70-1 and PfHsp70-z. PfHsp70-z (Hsp110 homologue) is thought to facilitate nucleotide exchange function of PfHsp70-1. PfHsp70-1 is a refoldase, while PfHsp70-z is restricted to holdase chaperone function. The structural features delineating functional specialisation of these chaperones remain unknown. Notably, PfHsp70-z possesses a unique linker segment which could account for its distinct functions. Using recombinant forms of PfHsp70-1, PfHsp70-z and E. coli Hsp70 (DnaK) as well as their linker switch mutant forms, we explored the effects of the linker mutations by conducting several assays such as circular dichroism, intrinsic and extrinsic fluorescence coupled to biochemical and in cellular analyses. Our findings demonstrate that the linker of PfHsp70-z modulates global conformation of the chaperone, regulating several functions such as client protein binding, chaperone- and ATPase activities. In addition, as opposed to the flexible linker of PfHsp70-1, the PfHsp70-z linker is rigid, thus regulating its notable thermal stability, making it an effective stress buffer. Our findings suggest a crucial role for the linker in streamlining the functions of these two chaperones. The findings further explain how these distinct chaperones cooperate to ensure survival of P. falciparum particularly under the stressful human host environment.

The Role of Non-Canonical Hsp70s (Hsp110/Grp170) in Cancer.

Although cancers account for over 16% of all global deaths annually, at present, no reliable therapies exist for most types of the disease. As protein folding facilitators, heat shock proteins (Hsps) play an important role in cancer development. Not surprisingly, Hsps are among leading anticancer drug targets. Generally, Hsp70s are divided into two main subtypes: canonical Hsp70 (Escherichia coli Hsp70/DnaK homologues) and the non-canonical (Hsp110 and Grp170) members. These two main Hsp70 groups are delineated from each other by distinct structural and functional specifications. Non-canonical Hsp70s are considered as holdase chaperones, while canonical Hsp70s are refoldases. This unique characteristic feature is mirrored by the distinct structural features of these two groups of chaperones. Hsp110/Grp170 members are larger as they possess an extended acidic insertion in their substrate binding domains. While the role of canonical Hsp70s in cancer has received a fair share of attention, the roles of non-canonical Hsp70s in cancer development has received less attention in comparison. In the current review, we discuss the structure-function features of non-canonical Hsp70s members and how these features impact their role in cancer development. We further mapped out their interactome and discussed the prospects of targeting these proteins in cancer therapy.

Comparative Characterization of Plasmodium falciparum Hsp70-1 Relative to E. coli DnaK Reveals the Functional Specificity of the Parasite Chaperone.

Hsp70 is a conserved molecular chaperone. How Hsp70 exhibits specialized functions across species remains to be understood. Plasmodium falciparum Hsp70-1 (PfHsp70-1) and Escherichia coli DnaK are cytosol localized molecular chaperones that are important for the survival of these two organisms. In the current study, we investigated comparative structure-function features of PfHsp70-1 relative to DnaK and a chimeric protein, KPf, constituted by the ATPase domain of DnaK and the substrate binding domain (SBD) of PfHsp70-1. Recombinant forms of the three Hsp70s exhibited similar secondary and tertiary structural folds. However, compared to DnaK, both KPf and PfHsp70-1 were more stable to heat stress and exhibited higher basal ATPase activity. In addition, PfHsp70-1 preferentially bound to asparagine rich peptide substrates, as opposed to DnaK. Recombinant P. falciparum adenosylmethionine decarboxylase (PfAdoMetDC) co-expressed in E. coli with either KPf or PfHsp70-1 was produced as a fully folded product. Co-expression of PfAdoMetDC with heterologous DnaK in E. coli did not promote folding of the former. However, a combination of supplementary GroEL plus DnaK improved folding of PfAdoMetDC. These findings demonstrated that the SBD of PfHsp70-1 regulates several functional features of the protein and that this molecular chaperone is tailored to facilitate folding of plasmodial proteins.

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.

The Link That Binds: The Linker of Hsp70 as a Helm of the Protein's Function.

The heat shock 70 (Hsp70) family of molecular chaperones plays a central role in maintaining cellular proteostasis. Structurally, Hsp70s are composed of an N-terminal nucleotide binding domain (NBD) which exhibits ATPase activity, and a C-terminal substrate binding domain (SBD). The binding of ATP at the NBD and its subsequent hydrolysis influences the substrate binding affinity of the SBD through allostery. Similarly, peptide binding at the C-terminal SBD stimulates ATP hydrolysis by the N-terminal NBD. Interdomain communication between the NBD and SBD is facilitated by a conserved linker segment. Hsp70s form two main subgroups. Canonical Hsp70 members generally suppress protein aggregation and are also capable of refolding misfolded proteins. Hsp110 members are characterized by an extended lid segment and their function tends to be largely restricted to suppression of protein aggregation. In addition, the latter serve as nucleotide exchange factors (NEFs) of canonical Hsp70s. The linker of the Hsp110 family is less conserved compared to that of the canonical Hsp70 group. In addition, the linker plays a crucial role in defining the functional features of these two groups of Hsp70. Generally, the linker of Hsp70 is quite small and varies in size from seven to thirteen residues. Due to its small size, any sequence variation that Hsp70 exhibits in this motif has a major and unique influence on the function of the protein. Based on sequence data, we observed that canonical Hsp70s possess a linker that is distinct from similar segments present in Hsp110 proteins. In addition, Hsp110 linker motifs from various genera are distinct suggesting that their unique features regulate the flexibility with which the NBD and SBD of these proteins communicate via allostery. The Hsp70 linker modulates various structure-function features of Hsp70 such as its global conformation, affinity for peptide substrate and interaction with co-chaperones. The current review discusses how the unique features of the Hsp70 linker accounts for the functional specialization of this group of molecular chaperones.

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.

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.

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.

Use of a Chimeric Hsp70 to Enhance the Quality of Recombinant Plasmodium falciparum S-Adenosylmethionine Decarboxylase Protein Produced in Escherichia coli.

S-adenosylmethionine decarboxylase (PfAdoMetDC) from Plasmodium falciparum is a prospective antimalarial drug target. The production of recombinant PfAdoMetDC for biochemical validation as a drug target is important. The production of PfAdoMetDC in Escherichia coli has been reported to result in unsatisfactory yields and poor quality product. The co-expression of recombinant proteins with molecular chaperones has been proposed as one way to improve the production of the former in E. coli. E. coli heat shock proteins DnaK, GroEL-GroES and DnaJ have previously been used to enhance production of some recombinant proteins. However, the outcomes were inconsistent. An Hsp70 chimeric protein, KPf, which is made up of the ATPase domain of E. coli DnaK and the substrate binding domain of P. falciparum Hsp70 (PfHsp70) has been previously shown to exhibit chaperone function when it was expressed in E. coli cells whose resident Hsp70 (DnaK) function was impaired. We proposed that because of its domain constitution, KPf would most likely be recognised by E. coli Hsp70 co-chaperones. Furthermore, because it possesses a substrate binding domain of plasmodial origin, KPf would be primed to recognise recombinant PfAdoMetDC expressed in E. coli. First, using site-directed mutagenesis, followed by complementation assays, we established that KPf with a mutation in the hydrophobic residue located in its substrate binding cavity was functionally compromised. We further co-expressed PfAdoMetDC with KPf, PfHsp70 and DnaK in E. coli cells either in the absence or presence of over-expressed GroEL-GroES chaperonin. The folded and functional status of the produced PfAdoMetDC was assessed using limited proteolysis and enzyme assays. PfAdoMetDC co-expressed with KPf and PfHsp70 exhibited improved activity compared to protein co-expressed with over-expressed DnaK. Our findings suggest that chimeric KPf may be an ideal Hsp70 co-expression partner for the production of recombinant plasmodial proteins in E. coli.

Plasmodium falciparum Hsp70-z, an Hsp110 homologue, exhibits independent chaperone activity and interacts with Hsp70-1 in a nucleotide-dependent fashion.

The role of molecular chaperones, among them heat shock proteins (Hsps), in the development of malaria parasites has been well documented. Hsp70s are molecular chaperones that facilitate protein folding. Hsp70 proteins are composed of an N-terminal nucleotide binding domain (NBD), which confers them with ATPase activity and a C-terminal substrate binding domain (SBD). In the ADP-bound state, Hsp70 possesses high affinity for substrate and releases the folded substrate when it is bound to ATP. The two domains are connected by a conserved linker segment. Hsp110 proteins possess an extended lid segment, a feature that distinguishes them from canonical Hsp70s. Plasmodium falciparum Hsp70-z (PfHsp70-z) is a member of the Hsp110 family of Hsp70-like proteins. PfHsp70-z is essential for survival of malaria parasites and is thought to play an important role as a molecular chaperone and nucleotide exchange factor of its cytosolic canonical Hsp70 counterpart, PfHsp70-1. Unlike PfHsp70-1 whose functions are fairly well established, the structure-function features of PfHsp70-z remain to be fully elucidated. In the current study, we established that PfHsp70-z possesses independent chaperone activity. In fact, PfHsp70-z appears to be marginally more effective in suppressing protein aggregation than its cytosol-localized partner, PfHsp70-1. Furthermore, based on coimmunoaffinity chromatography and surface plasmon resonance analyses, PfHsp70-z associated with PfHsp70-1 in a nucleotide-dependent fashion. Our findings suggest that besides serving as a molecular chaperone, PfHsp70-z could facilitate the nucleotide exchange function of PfHsp70-1. These dual functions explain why it is essential for parasite survival.

Overexpression, Purification and Characterisation of the Plasmodium falciparum Hsp70-z (PfHsp70-z) Protein.

Six Hsp70-like genes are represented on the genome of Plasmodium falciparum. Of these two occur in the cytosol: P. falciparum Hsp70-z (PfHsp70-z) and PfHsp70-1. PfHsp70-1 is a well characterised canonical Hsp70 that facilitates protein quality control and is crucial for the development of malaria parasites. There is very little known about PfHsp70-z. However, PfHsp70-z is known to be essential and is implicated in suppressing aggregation of asparagine-rich proteins of P. falciparum. In addition, its expression at the clinical stage of malaria correlates with disease prognosis. Based on structural evidence PfHsp70-z belongs to the Hsp110 family of proteins. Since Hsp110 proteins have been described as nucleotide exchange factors (NEFs) of their canonical Hsp70 counterparts, it has been speculated that PfHsp70-z may serve as a NEF of PfHsp70-1. In the current study, P. falciparum cells cultured in vitro were subjected to heat stress, triggering the enhanced expression of PfHsp70-z. Biochemical assays conducted using recombinant PfHsp70-z protein demonstrated that the protein is heat stable and possesses ATPase activity. Furthermore, we observed that PfHsp70-z is capable of self-association. The structural-functional features of PfHsp70-z provide further evidence for its role as a chaperone and possible nucleotide exchange factor of PfHsp70-1.

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.

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.

An in vitro assessment of the interaction of cadmium selenide quantum dots with DNA, iron, and blood platelets.

Cadmium selenide (CdSe) quantum dots have gained increased attention for their potential use in biomedical applications. This has raised interest in assessing their toxicity. In this study, water-soluble, cysteine-capped CdSe nanocrystals with an average size of 15 nm were prepared through a one-pot solution-based method. The CdSe nanoparticles were synthesized in batches in which the concentration of the capping agent was varied with the aim of stabilizing the quantum dot core. The effects of the CdSe quantum dots on DNA stability, aggregation of blood platelets, and reducing activity of iron were evaluated in vitro . DNA damage was observed at a concentration of 200 µg/mL of CdSe quantum dots. Furthermore, the CdSe nanocrystals exhibited high reducing power and chelating activity, suggesting that they may impair the function of haemoglobin by interacting with iron. In addition, the CdSe quantum dots promoted aggregation of blood platelets in a dose dependent manner.

Plasmodium falciparum encodes a single cytosolic type I Hsp40 that functionally interacts with Hsp70 and is upregulated by heat shock.

Heat shock protein 70 (Hsp70) and heat shock protein 40 (Hsp40) function as molecular chaperones during the folding and trafficking of proteins within most cell types. However, the Hsp70-Hsp40 chaperone partnerships within the malaria parasite, Plasmodium falciparum, have not been elucidated. Only one of the 43 P. falciparum Hsp40s is predicted to be a cytosolic, canonical Hsp40 (termed PfHsp40) capable of interacting with the major cytosolic P. falciparum-encoded Hsp70, PfHsp70. Consistent with this hypothesis, we found that PfHsp40 is upregulated under heat shock conditions in a similar pattern to PfHsp70. In addition, PfHsp70 and PfHsp40 reside mainly in the parasite cytosol, as assessed using indirect immunofluorescence microscopy. Recombinant PfHsp40 stimulated the ATP hydrolytic rates of both PfHsp70 and human Hsp70 similar to other canonical Hsp40s of yeast (Ydj1) and human (Hdj2) origin. In contrast, the Hsp40-stimulated plasmodial and human Hsp70 ATPase activities were differentially inhibited in the presence of pyrimidinone-based small molecule modulators. To further probe the chaperone properties of PfHsp40, protein aggregation suppression assays were conducted. PfHsp40 alone suppressed protein aggregation, and cooperated with PfHsp70 to suppress aggregation. Together, these data represent the first cellular and biochemical evidence for a PfHsp70-PfHsp40 partnership in the malaria parasite, and furthermore that the plasmodial and human Hsp70-Hsp40 chaperones possess unique attributes that are differentially modulated by small molecules.