Plasmodium falciparum is the most lethal of human-infective malaria parasites. A hallmark of P. falciparum malaria is extensive remodeling of host erythrocytes by the parasite, which facilitates the development of virulence properties such as host cell adhesion to the endothelial lining of the microvasculature. Host remodeling is mediated by a large complement of parasite proteins exported to the erythrocyte; among them is a single heat shock protein (Hsp)70–class protein chaperone, P. falciparum Hsp70-x (PfHsp70-x). PfHsp70-x was previously shown to assist the development of virulent cytoadherence characteristics. Here, we show that PfHsp70-x also supports parasite growth under elevated temperature conditions that simulate febrile episodes, especially at the beginning of the parasite life cycle when most of host cell remodeling takes place. Biochemical and biophysical analyses of PfHsp70-x, including crystallographic structures of its catalytic domain and the J-domain of its stimulatory Hsp40 cochaperone, suggest that PfHsp70-x is highly similar to human Hsp70 chaperones endogenous to the erythrocyte. Nevertheless, our results indicate that selective inhibition of PfHsp70-x function using small molecules may be possible and highlight specific sites of its catalytic domain as potentially of high interest. We discuss the likely roles of PfHsp70-x and human chaperones in P. falciparum biology and how specific inhibitors may assist us in disentangling their relative contributions.—Day, J., Passecker, A., Beck, H.-P., Vakonakis, I. The Plasmodium falciparum Hsp70-x chaperone assists the heat stress response of the malaria parasite.

Native mass spectrometry (MS) methods permit the study of multiple protein species within solution equilibria, whereas ion mobility (IM)-MS can report on conformational behavior of specific states. We used IM-MS to study a conformationally labile protein (α1-antitrypsin) that undergoes pathological polymerization in the context of point mutations. The folded, native state of the Z-variant remains highly polymerogenic in physiological conditions despite only minor thermodynamic destabilization relative to the wild-type variant. Various data implicate kinetic instability (conformational lability within a native state ensemble) as the basis of Z α1-antitrypsin polymerogenicity. We show the ability of IM-MS to track such disease-relevant conformational behavior in detail by studying the effects of peptide binding on α1-antitrypsin conformation and dynamics. IM-MS is, therefore, an ideal platform for the screening of compounds that result in therapeutically beneficial kinetic stabilization of native α1-antitrypsin. Our findings are confirmed with high-resolution X-ray crystallographic and nuclear magnetic resonance spectroscopic studies of the same event, which together dissect structural changes from dynamic effects caused by peptide binding at a residue-specific level. IM-MS methods, therefore, have great potential for further study of biologically relevant thermodynamic and kinetic instability of proteins and provide rapid and multidimensional characterization of ligand interactions of therapeutic interest.PDB Code(s): 4PYW

SummaryAdherence of Plasmodium falciparum‐infected erythrocytes to host endothelium is conferred through the parasite‐derived virulence factor P. falciparum erythrocyte membrane protein 1 (PfEMP1), the major contributor to malaria severity. PfEMP1 located at knob structures on the erythrocyte surface is anchored to the cytoskeleton, and the Plasmodium helical interspersed subtelomeric (PHIST) gene family plays a role in many host cell modifications including binding the intracellular domain of PfEMP1. Here, we show that conditional reduction of the PHIST protein PFE1605w strongly reduces adhesion of infected erythrocytes to the endothelial receptor CD36. Adhesion to other endothelial receptors was less affected or even unaltered by PFE1605w depletion, suggesting that PHIST proteins might be optimized for subsets of PfEMP1 variants. PFE1605w does not play a role in PfEMP1 transport, but it directly interacts with both the intracellular segment of PfEMP1 and with cytoskeletal components. This is the first report of a PHIST protein interacting with key molecules of the cytoadherence complex and the host cytoskeleton, and this functional role seems to play an essential role in the pathology of P. falciparum.

No abstract