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Structure of anthrax lethal toxin prepore complex suggests a pathway for efficient cell entry

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ABSTRACT

Anthrax toxin is a tripartite complex in which the protective antigen moiety forms a pore through which lethal factor and edema factor are translocated. Fabre et al. reveal a mechanism for efficient translocation in their structure of the heptameric protective antigen prepore bound to three lethal factors.

No MeSH data available.


Related in: MedlinePlus

3-D map of the (PA63)7–(LF)3 complex and its atomic interpretation. (A–D) The map of (PA63)7–(LF)3 obtained by cryo-EM single-particle reconstruction at 16-Å resolution is shown in isosurface representation. (A) Side view showing the upside down L shape of LF molecules with a gap of ∼30 Å tall (blue oval) between the surface of PA and the LFC domains. (B) Side view, rotated about a vertical axis by ∼100° counterclockwise compared with A, showing the major intermolecular LFC–LFN interaction within the complex (circled in red here and in C), and a minor contact visible at lower contour levels (red arrows here and in C). (C) Top-down view of the complex showing three LF molecules, each in unique environment (named from above 1LF, 2LF, and 3LF; see F), Note the “gap” (marked by black arrow) caused by the symmetry mismatch, the major intermolecular LFC–LFN (circled in red), and the minor contact visible at a lower contour level (red arrows). (D) View from below reveals the strong sevenfold symmetry of (PA63)7, which was not imposed during refinement. (E and F) Atomic models derived by rigid-body docking of component crystal structures into the EM map. The view in E looks directly into the gap, which overlooks the orphan PA63 moiety and is a rotation of A by 100° clockwise. The view in F is the same as C. The three LF molecules fit their respective densities with CC > 0.98, whereas the junctions between the LF molecules and their cognate PA molecule have CC > 0.97. Bars, 50 Å.
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fig4: 3-D map of the (PA63)7–(LF)3 complex and its atomic interpretation. (A–D) The map of (PA63)7–(LF)3 obtained by cryo-EM single-particle reconstruction at 16-Å resolution is shown in isosurface representation. (A) Side view showing the upside down L shape of LF molecules with a gap of ∼30 Å tall (blue oval) between the surface of PA and the LFC domains. (B) Side view, rotated about a vertical axis by ∼100° counterclockwise compared with A, showing the major intermolecular LFC–LFN interaction within the complex (circled in red here and in C), and a minor contact visible at lower contour levels (red arrows here and in C). (C) Top-down view of the complex showing three LF molecules, each in unique environment (named from above 1LF, 2LF, and 3LF; see F), Note the “gap” (marked by black arrow) caused by the symmetry mismatch, the major intermolecular LFC–LFN (circled in red), and the minor contact visible at a lower contour level (red arrows). (D) View from below reveals the strong sevenfold symmetry of (PA63)7, which was not imposed during refinement. (E and F) Atomic models derived by rigid-body docking of component crystal structures into the EM map. The view in E looks directly into the gap, which overlooks the orphan PA63 moiety and is a rotation of A by 100° clockwise. The view in F is the same as C. The three LF molecules fit their respective densities with CC > 0.98, whereas the junctions between the LF molecules and their cognate PA molecule have CC > 0.97. Bars, 50 Å.

Mentions: We assembled such a complex in vitro by mixing preformed (PA63)7 heptamers with LF, which produced a (PA63)7–(LF)3 complex with the expected stoichiometry (Cunningham et al., 2002; Mogridge et al., 2002), as judged by gel filtration and SDS page (Fig. 1). We then determined the structure of this complex at neutral pH in vitreous ice to 16-Å resolution using single-particle cryo-EM methods (Figs. 1, 2, and 3; and Figs. S1 and S2). Our map bears little resemblance to the previously published (PA63)7–(LF)1 structure. Instead, it displays a highly symmetric (sevenfold) core, with three large inverted L-shaped densities on top (Fig. 4). Initial rigid-body fitting of crystal structures of the (PA63)7 prepore (Lacy et al., 2004) into the symmetric core and three full-length (monomeric) LF molecules (Pannifer et al., 2001) into the L-shaped densities gave excellent fits that accounted for most of the density (overall correlation coefficient [CC] = 0.94). The conformations of the three LF molecules and their interactions with PA protomers are nearly identical at this resolution (Fig. 4). We also note that no quaternary changes within LF (e.g., hinge bending between the LFN and LFC) were observed; indeed, the three densities were identical with a cross-correlation >0.98. Image analysis showed the absence of octamers in our assembly conditions.


Structure of anthrax lethal toxin prepore complex suggests a pathway for efficient cell entry
3-D map of the (PA63)7–(LF)3 complex and its atomic interpretation. (A–D) The map of (PA63)7–(LF)3 obtained by cryo-EM single-particle reconstruction at 16-Å resolution is shown in isosurface representation. (A) Side view showing the upside down L shape of LF molecules with a gap of ∼30 Å tall (blue oval) between the surface of PA and the LFC domains. (B) Side view, rotated about a vertical axis by ∼100° counterclockwise compared with A, showing the major intermolecular LFC–LFN interaction within the complex (circled in red here and in C), and a minor contact visible at lower contour levels (red arrows here and in C). (C) Top-down view of the complex showing three LF molecules, each in unique environment (named from above 1LF, 2LF, and 3LF; see F), Note the “gap” (marked by black arrow) caused by the symmetry mismatch, the major intermolecular LFC–LFN (circled in red), and the minor contact visible at a lower contour level (red arrows). (D) View from below reveals the strong sevenfold symmetry of (PA63)7, which was not imposed during refinement. (E and F) Atomic models derived by rigid-body docking of component crystal structures into the EM map. The view in E looks directly into the gap, which overlooks the orphan PA63 moiety and is a rotation of A by 100° clockwise. The view in F is the same as C. The three LF molecules fit their respective densities with CC > 0.98, whereas the junctions between the LF molecules and their cognate PA molecule have CC > 0.97. Bars, 50 Å.
© Copyright Policy - openaccess
Related In: Results  -  Collection

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fig4: 3-D map of the (PA63)7–(LF)3 complex and its atomic interpretation. (A–D) The map of (PA63)7–(LF)3 obtained by cryo-EM single-particle reconstruction at 16-Å resolution is shown in isosurface representation. (A) Side view showing the upside down L shape of LF molecules with a gap of ∼30 Å tall (blue oval) between the surface of PA and the LFC domains. (B) Side view, rotated about a vertical axis by ∼100° counterclockwise compared with A, showing the major intermolecular LFC–LFN interaction within the complex (circled in red here and in C), and a minor contact visible at lower contour levels (red arrows here and in C). (C) Top-down view of the complex showing three LF molecules, each in unique environment (named from above 1LF, 2LF, and 3LF; see F), Note the “gap” (marked by black arrow) caused by the symmetry mismatch, the major intermolecular LFC–LFN (circled in red), and the minor contact visible at a lower contour level (red arrows). (D) View from below reveals the strong sevenfold symmetry of (PA63)7, which was not imposed during refinement. (E and F) Atomic models derived by rigid-body docking of component crystal structures into the EM map. The view in E looks directly into the gap, which overlooks the orphan PA63 moiety and is a rotation of A by 100° clockwise. The view in F is the same as C. The three LF molecules fit their respective densities with CC > 0.98, whereas the junctions between the LF molecules and their cognate PA molecule have CC > 0.97. Bars, 50 Å.
Mentions: We assembled such a complex in vitro by mixing preformed (PA63)7 heptamers with LF, which produced a (PA63)7–(LF)3 complex with the expected stoichiometry (Cunningham et al., 2002; Mogridge et al., 2002), as judged by gel filtration and SDS page (Fig. 1). We then determined the structure of this complex at neutral pH in vitreous ice to 16-Å resolution using single-particle cryo-EM methods (Figs. 1, 2, and 3; and Figs. S1 and S2). Our map bears little resemblance to the previously published (PA63)7–(LF)1 structure. Instead, it displays a highly symmetric (sevenfold) core, with three large inverted L-shaped densities on top (Fig. 4). Initial rigid-body fitting of crystal structures of the (PA63)7 prepore (Lacy et al., 2004) into the symmetric core and three full-length (monomeric) LF molecules (Pannifer et al., 2001) into the L-shaped densities gave excellent fits that accounted for most of the density (overall correlation coefficient [CC] = 0.94). The conformations of the three LF molecules and their interactions with PA protomers are nearly identical at this resolution (Fig. 4). We also note that no quaternary changes within LF (e.g., hinge bending between the LFN and LFC) were observed; indeed, the three densities were identical with a cross-correlation >0.98. Image analysis showed the absence of octamers in our assembly conditions.

View Article: PubMed Central - HTML - PubMed

ABSTRACT

Anthrax toxin is a tripartite complex in which the protective antigen moiety forms a pore through which lethal factor and edema factor are translocated. Fabre et al. reveal a mechanism for efficient translocation in their structure of the heptameric protective antigen prepore bound to three lethal factors.

No MeSH data available.


Related in: MedlinePlus