Structural–functional diversity of malaria parasite's PfHSP70-1 and PfHSP40 chaperone pair gives an edge over human orthologs in chaperone-assisted protein folding

Abstract

Plasmodium falciparum, the human malaria parasite harbors a metastable proteome which is vulnerable to proteotoxic stress conditions encountered during its lifecycle. How parasite’s chaperone machinery is able to maintain its aggregation-prone proteome in functional state, is poorly understood. As HSP70-40 system forms the central hub in cellular proteostasis, we investigated the protein folding capacity of PfHSP70-1 and PfHSP40 chaperone pair and compared it with human orthologs (HSPA1A and DNAJA1). Despite structural similarity, we observed that parasite chaperones and their human orthologs exhibit striking differences in conformational dynamics. Comprehensive biochemical investigations revealed that PfHSP70-1 and PfHSP40 chaperone pair has better protein folding, aggregation inhibition and oligomer remodeling and disaggregase activities than their human orthologs. Chaperone-swapping experiments suggest that PfHSP40 can also efficiently cooperate with human HSP70 to facilitate folding of client-substrate. SPR-derived kinetic parameters reveal that PfHSP40 has higher binding affinity towards unfolded substrate than DNAJA1. Interestingly, the observed slow dissociation rate of PfHSP40-substrate interaction allows PfHSP40 to maintain substrate in folding-competent state to minimize its misfolding. Structural investigation through SAXS gave insights into the conformational architecture of PfHSP70-1 (monomer), PfHSP40 (dimer) and their complex. Overall, our data suggests that parasite has evolved functionally diverged and efficient chaperone machinery which allows human malaria parasite to survive in hostile conditions. The distinct allosteric landscapes and interaction kinetics of plasmodial chaperones open avenues for exploration of small-molecule based antimalarial interventions.

Adenine Modification at C7 as a Viable Strategy to Potentiate the Antimalarial Activity of Quinolones

“…For luciferin-luciferase assay, the luciferase was purified as described previously. [53] The assay buffer composition was 50 mM Tris pH 7.5, 12 mM MgCl 2 , 0.3 μM luciferase, 1 mM luciferin). Luminescence was recorded in FLX800, BIOTEK and was normalized with respective protein concentration.…”

Plasmodium falciparum Molecular Chaperones: Guardians of the Malaria Parasite Proteome and Renovators of the Host Proteome

“…The cytoplasmic PfHsp70-z is also essential ( Muralidharan et al, 2012 ; Zhang et al, 2018 ), which may well be due to its highly effective protein aggregation suppression activity ( Zininga et al, 2016 ). PfHsp70-1 has also been shown to have high ATPase activity ( Matambo et al, 2004 ; Misra and Ramachandran, 2009 ; Makumire et al, 2021 ) and strong aggregation suppression activity ( Botha et al, 2011 ), suggesting that it is a superior chaperone compared to human Hsp70s ( Anas et al, 2020 ). Overall, the evidence suggests that PfHsp70-1 and PfHsp70-z are major players in proteostasis of the parasite cytoplasm.…”

“…A number of the parasite-resident PfJDPs, have been identified as potential co-chaperone of PfHsp70-1 (PfHsp40/PF14_0359, Botha et al, 2011 , Anas et al, 2020 ; PFB0595w, Njunge et al, 2015 ; and Pfj4/PFL0565w, Pesce et al, 2008 ). Like PfHsp70-1, PfHsp40 is essential ( Zhang et al, 2018 ), cytosolic, constitutively expressed, and upregulated under stressful conditions ( Botha et al, 2011 ).…”

“…Like PfHsp70-1, PfHsp40 is essential ( Zhang et al, 2018 ), cytosolic, constitutively expressed, and upregulated under stressful conditions ( Botha et al, 2011 ). While they are the homologous chaperone pair to the canonical cytosolic human Hsp70-JDP pair (HSPA1A-DNAJA1), there are subtle but critical structural, biochemical and functional differences, with the P. falciparum pair shown to be a more effective chaperone machine ( Anas et al, 2020 ). Furthermore, PfHsp40 has been found to be farnesylated and palmitoylated, leading to membrane localization ( Mathews et al, 2021 ).…”

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

“…However, targeting P. falciparum Hsps is a challenge, as these proteins are generally conserved ( Pesce et al, 2010 ; Chakafana et al, 2019 ). Despite this, there is growing evidence that Hsp70s of parasitic origin, and in particular PfHsp70-1, exhibit distinct structure-function features that could make them amenable to selective targeting ( Chakafana et al, 2019 ; Anas et al, 2020 ; Lebepe et al, 2020 ; Makumire et al, 2021 ). Promisingly, some antimalarial compounds demonstrating selective inhibition of parasite Hsp70 with minimum effects on the function of human Hsp70 have been described ( Cockburn et al, 2014 ; reviewed in Zininga and Shonhai, 2019 ).…”