Malaria Parasite Actin Polymerization and Filament Structure

Stephan Schmitz, Iwan A. T. Schaap, Jens Kleinjung, Simone Harder, Munira Grainger, Lesley Calder, Peter B. Rosenthal, Anthony A. Holder, Claudia Veigel

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Abstract

A novel form of acto-myosin regulation has been proposed in which polymerization of new actin filaments regulates motility of parasites of the apicomplexan class of protozoa. In vivo and in vitro parasite F-actin is very short and unstable, but the structural basis and details of filament dynamics remain unknown. Here, we show that long actin filaments can be obtained by polymerizing unlabeled rabbit skeletal actin (RS-actin) onto both ends of the short rhodamine-phalloidin-stabilized Plasmodium falciparum actin I (Pf-actin) filaments. Following annealing, hybrid filaments of micron length and "zebra-striped" appearance are observed by fluorescence microscopy that are stable enough to move over myosin class II motors in a gliding filament assay. Using negative stain electron microscopy we find that pure Pf-actin stabilized by jasplakinolide (JAS) also forms long filaments, indistinguishable in length from RS-actin filaments, and long enough to be characterized structurally. To compare structures in near physiological conditions in aqueous solution we imaged Pf-actin and RS-actin filaments by atomic force microscopy (AFM). We found the monomer stacking to be distinctly different for Pf-actin compared with RS-actin, such that the pitch of the double helix of Pf-actin filaments was 10% larger. Our results can be explained by a rotational angle between subunits that is larger in the parasite compared with RS-actin. Modeling of the AFM data using high-resolution actin filament models supports our interpretation of the data. The structural differences reported here may be a consequence of weaker inter-and intra-strand contacts, and may be critical for differences in filament dynamics and for regulation of parasite motility.

Original languageEnglish
Pages (from-to)36577-36585
Number of pages9
JournalJournal of Biological Chemistry
Volume285
Issue number47
DOIs
Publication statusPublished - 19 Nov 2010

Keywords

  • PLASMODIUM-FALCIPARUM ACTIN
  • RAY FIBER DIFFRACTION
  • F-ACTIN
  • TOXOPLASMA-GONDII
  • APICOMPLEXAN PARASITES
  • ADP-ACTIN
  • FLUORESCENCE MICROSCOPY
  • MICROFILAMENT DYNAMICS
  • GLIDING MOTILITY
  • BINDING PROTEINS

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