Biochemical and biophysical characterization of Plasmodium falciparum glucose regulated protein 170

The main malaria parasite, Plasmodium falciparum expresses 6 Hsp70 proteins. Among them, two are Hsp110 homologues localised in the cytosol and endoplasmic reticulum. The cytosolic Hsp110 (PfHsp70-z) has been fairly studied but the ER, PfGrp170 (PfHsp70-y) has not been well characterised. PfGrp170 is essential for parasite survival and is thought to be important for protein folding during exportation of parasite proteins to the host cell cytosol. Therefore, this study optimised the conditions for its overexpression in an E. coli system and investigated the structure-functional features of Grp170 protein. The recombinant full-length version of PfGrp170 was successfully produced under native conditions. The secondary structure analysis showed a predominantly α-helical structural features which exhibit moderate thermal stability with a Tm above 55 °C. The presence of nucleotides, such as ATP perturbed the structural stability of PfGrp170 making it more sensitive to heat stress and proteolytic cleavage. We observed that PfGrp170 exhibits basal ATPase activity and has the capability to suppress the heat induced aggregation of model substrate malate dehydrogenase in a nucleotide independent manner. In addition, using molecular docking and molecular dynamics simulations we show evidence of selective binding to nucleotides and the protein structure is more responsive to ATP binding than ADP. Findings from this study provide the first direct evidence for the independent chaperone function of PfGrp170 and its sensitivity to structural modulation by nucleotides which could be targeted in antimalarial drug development strategies.