Establishing Computational Approaches Towards Identifying Malarial Allosteric Modulators: A Case Study of Plasmodium falciparum Hsp70s

Abstract

Combating malaria is almost a never-ending battle, as Plasmodium parasites develop resistance to the drugs used against them, as observed recently in artemisinin-based combination therapies. The main concern now is if the resistant parasite strains spread from Southeast Asia to Africa, the continent hosting most malaria cases. To prevent catastrophic results, we need to find non-conventional approaches. Allosteric drug targeting sites and modulators might be a new hope for malarial treatments. Heat shock proteins (HSPs) are potential malarial drug targets and have complex allosteric control mechanisms. Yet, studies on designing allosteric modulators against them are limited. Here, we identified allosteric modulators (SANC190 and SANC651) against P. falciparum Hsp70-1 and Hsp70-x, affecting the conformational dynamics of the proteins, delicately balanced by the endogenous ligands. Previously, we established a pipeline to identify allosteric sites and modulators. This study also further investigated alternative approaches to speed up the process by comparing all atom molecular dynamics simulations and dynamic residue network analysis with the coarse-grained (CG) versions of the calculations. Betweenness centrality (BC) profiles for PfHsp70-1 and PfHsp70-x derived from CG simulations not only revealed similar trends but also pointed to the same functional regions and specific residues corresponding to BC profile peaks.

Understanding the Pyrimethamine Drug Resistance Mechanism via Combined Molecular Dynamics and Dynamic Residue Network Analysis

“…Intriguingly, some of the residues possessing high BC values, 10-14, 57-63, 159-162, and 180-184, form part of the positively charged groove at the back of the active site responsible for critical interactions (with the negatively charged junction region helix 285-294) necessary for Pf DHFR-TS protein folding and function [21,34,35]. These findings are in agreement with previous studies which found high centrality residues around active site and linker interphases of protein domains [23,30,33]. Using WT holoenzyme as reference, twenty-four key communication residues were identified: 10, 13, 15, 16, 18, 21, 41, 55, 59, 63, 101, 103, 105, 109, 159, 162, 163, 165, 167, 170, 180, 181, 185, and 196.…”

“…Overall, regions showing significantly high BC values were located within or close to substrate/ligand-binding sites (shaded grey in Figure 7). These results recapitulate previous findings reporting that active sites of proteins possess high and low average BC and average L values, respectively [23,30,33].…”

“…To be able to observe potential discrete conformational changes in protein systems and hence to understand the variation effect on them, RMSD distribution histograms of the WT and variant proteins both in ligand free and ligand bound were generated, as previously applied [22][23][24] (Figure 2A). The pyrimethamine-free WT model displayed nearly normal RMSD distribution while the bound system showed a bimodal curve, suggesting that the former predominantly sampled a single conformer while the latter sampled at least two conformations during simulation.…”

“…Both metrics accentuate residue importance in protein communication. The L and BC measures become even more robust when computed as running averages across an MD trajectory, in what is known as dynamic residue network (DRN) analysis [23,24,30,32]. We present per residue reachability as plots of average L in (Figure 6).…”

Integrated Computational Approaches and Tools for Allosteric Drug Discovery

“…Another important incentive for the development of allosteric drugs is that, while traditional orthosteric drugs usually inhibit protein activity, allosteric modulators may not only inhibit but also increase protein activity (allosteric activators) [29]. In the last decade, drug discovery has been shifting its focus toward targeting allosteric sites in order to improve compound selectivity [28][29][30][31][32][33]. Allosteric drugs also feature distinct physicochemical properties, adding further freedom for discovery of novel active compounds, and can often be combined with orthosteric drugs into synergistic drug cocktails to modulate and improve enzyme activities, specificity, and pharmacological profiles.…”