pH-dependence of the Plasmodium falciparum chloroquine resistance transporter is linked to the transport cycle

• Published in: Nature communications 2023-07, Vol.14 (1), p.4234-4234, Article 4234
• Main Authors: Berger, Fiona, Gomez, Guillermo M., Sanchez, Cecilia P., Posch, Britta, Planelles, Gabrielle, Sohraby, Farzin, Nunes-Alves, Ariane, Lanzer, Michael
• Format: Article
• Language: English
• Subjects: 38; 38/39; 38/70; 631/326/417/1716; 631/535/1267; 692/420/254; Alanine; Allosteric properties; Amino acids; Antimalarials - chemistry; Antimalarials - pharmacology; Carboxyl group; Chloroquine; Chloroquine - pharmacology; Detoxification; Drug Resistance - genetics; Humanities and Social Sciences; Humans; Hydrogen-Ion Concentration; Life Sciences; Malaria; Malaria, Falciparum - drug therapy; Malaria, Falciparum - parasitology; Membrane Transport Proteins - metabolism; Molecular dynamics; multidisciplinary; Mutagenesis; Parasite resistance; Parasites; pH effects; Plasmodium falciparum; Plasmodium falciparum - genetics; Plasmodium falciparum - metabolism; Proteolysis; Protons; Protozoa; Protozoan Proteins - metabolism; Residues; Scanning mutagenesis; Science; Science (multidisciplinary); Simulation; Substrate specificity; Substrates; Vector-borne diseases
• ISSN: 2041-1723; 2041-1723
• DOI: 10.1038/s41467-023-39969-2

The chloroquine resistance transporter, PfCRT, of the human malaria parasite Plasmodium falciparum is sensitive to acidic pH. Consequently, PfCRT operates at 60% of its maximal drug transport activity at the pH of 5.2 of the digestive vacuole, a proteolytic organelle from which PfCRT expels drugs interfering with heme detoxification. Here we show by alanine-scanning mutagenesis that E207 is critical for pH sensing. The E207A mutation abrogates pH-sensitivity, while preserving drug substrate specificity. Substituting E207 with Asp or His, but not other amino acids, restores pH-sensitivity. Molecular dynamics simulations and kinetics analyses suggest an allosteric binding model in which PfCRT can accept both protons and chloroquine in a partial noncompetitive manner, with increased proton concentrations decreasing drug transport. Further simulations reveal that E207 relocates from a peripheral to an engaged location during the transport cycle, forming a salt bridge with residue K80. We propose that the ionized carboxyl group of E207 acts as a hydrogen acceptor, facilitating transport cycle progression, with pH sensing as a by-product. PfCRT is a chloroquine resistance transporter from malaria parasite Plasmodium falciparum , which is sensitive to acidic pH. Here, the authors show that residue E207 is critical for pH sensing by PfCRT, using alanine-scanning mutagenesis, MD simulations and drug uptake assays.