Purification and initial characterization of Plasmodium falciparum K+ channels, PfKch1 and PfKch2 produced in Saccharomyces cerevisiae

Karen Molbaek, Maria Tejada, Christina Hoeier Ricke, Peter Scharff-Poulsen, Peter Ellekvist, Claus Helix-Nielsen, Nirbhay Kumar, Dan A. Klaerke, Per Amstrup Pedersen

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Abstract

Resistance towards known antimalarial drugs poses a significant problem, urging for novel drugs that target vital proteins in the malaria parasite Plasmodium falciparum. However, recombinant production of malaria proteins is notoriously difficult. To address this, we have investigated two putative K+ channels, PfKch1 and PfKch2, identified in the P. falciparum genome. We show that PfKch1 and PfKch2 and a C-terminally truncated version of PfKch1 (PfKch11-1094) could indeed be functionally expressed in vivo, since a K+-uptake deficient Saccharomyces cerevisiae strain was complemented by the P. falciparum cDNAs. PfKch11-1094-GFP and GFP-PfKch2 fusion proteins were overexpressed in yeast, purified and reconstituted in lipid bilayers to determine their electrophysiological activity. Single channel conductance amounted to 16 ± 1 pS for PfKch11-1094-GFP and 28 ± 2 pS for GFP-PfKch2. We predicted regulator of K+-conductance (RCK) domains in the C-terminals of both channels, and we accordingly measured channel activity in the presence of Ca2+.

Original languageEnglish
Article number183
JournalMicrobial Cell Factories
Volume19
Issue number1
Number of pages16
ISSN1475-2859
DOIs
Publication statusPublished - 2020

Keywords

  • K-channels
  • Malaria
  • Recombinant protein
  • Yeast

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