Fiber Formation across the Bacterial Outer Membrane by the Chaperone/Usher Pathway

Institute of Structural Molecular Biology, University College London and Birkbeck College, Malet Street, London, WC1E 7HX, United Kingdom.
Cell (Impact Factor: 32.24). 06/2008; 133(4):640-52. DOI: 10.1016/j.cell.2008.03.033
Source: PubMed


Gram-negative pathogens commonly exhibit adhesive pili on their surfaces that mediate specific attachment to the host. A major class of pili is assembled via the chaperone/usher pathway. Here, the structural basis for pilus fiber assembly and secretion performed by the outer membrane assembly platform--the usher--is revealed by the crystal structure of the translocation domain of the P pilus usher PapC and single particle cryo-electron microscopy imaging of the FimD usher bound to a translocating type 1 pilus assembly intermediate. These structures provide molecular snapshots of a twinned-pore translocation machinery in action. Unexpectedly, only one pore is used for secretion, while both usher protomers are used for chaperone-subunit complex recruitment. The translocating pore itself comprises 24 beta strands and is occluded by a folded plug domain, likely gated by a conformationally constrained beta-hairpin. These structures capture the secretion of a virulence factor across the outer membrane of gram-negative bacteria.

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    • "To illustrate guidance provided by the physical model, consider the mechanisms by which the chaperone-subunit complex of a diverse group of Gram-negative bacteria [28] gains passage through the Papc usher channel at the outer membrane. Here, an approaching chaperone complex triggers a response by which outer membrane usher passage unblocks. "
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    ABSTRACT: A diffusion theory-based, all-physical ab initio protein folding simulation is described and applied. The model is based upon the drift and diffusion of protein substructures relative to one another in the multiple energy fields present. Without templates or statistical inputs, the simulations were run at physiologic and ambient temperatures (including pH). Around 100 protein secondary structures were surveyed, and twenty tertiary structures were determined. Greater than 70% of the secondary core structures with over 80% alpha helices were correctly identified on protein ranging from 30 to 200 amino-acid sequence. The drift-diffusion model predicted tertiary structures with RMSD values in the 3-5 Angstroms range for proteins ranging 30 to 150 amino acids. These predictions are among the best for an all ab initio protein simulation. Simulations could be run entirely on a desktop computer in minutes; however, more accurate tertiary structures were obtained using molecular dynamic energy relaxation. The drift-diffusion model generated realistic energy versus time traces. Rapid secondary structures followed by a slow compacting towards lower energy tertiary structures occurred after an initial incubation period in agreement with observations.
    09/2013; 2013:583045. DOI:10.1155/2013/583045
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    • "Each subunit of these homopolymeric structures possesses the host-cell receptor binding site or sites; thus, they are polyadhesins. In contrast, the FGS chaperones assemble heteropolymeric, well-structured adhesive pili composed of up to seven different subunits [10,19]. Pili are monoadhesins, as they possess only one receptor binding subunit located at the tip of the organelle. "
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    ABSTRACT: Background The global spread of bacterial resistance has given rise to a growing interest in new anti-bacterial agents with a new strategy of action. Pilicides are derivatives of ring-fused 2-pyridones which block the formation of the pili/fimbriae crucial to bacterial pathogenesis. They impair by means of a chaperone-usher pathway conserved in the Gram-negative bacteria of adhesive structures biogenesis. Pili/fimbriae of this type belong to two subfamilies, FGS and FGL, which differ in the details of their assembly mechanism. The data published to date have shown that pilicides inhibit biogenesis of type 1 and P pili of the FGS type which are encoded by uropathogenic E. coli strains. Results We evaluated the anti-bacterial activity of literature pilicides as blockers of the assembly of a model example of FGL-type adhesive structures, – the Dr fimbriae encoded by a dra gene cluster of uropathogenic Escherichia coli strains. In comparison to the strain grown without pilicide, the Dr+ bacteria cultivated in the presence of the 3.5 mM concentration of pilicides resulted in a reduction of 75 to 87% in the adherence properties to CHO cells expressing Dr fimbrial DAF receptor protein. Using quantitative assays, we determined the amount of Dr fimbriae in the bacteria cultivated in the presence of 3.5 mM of pilicides to be reduced by 75 to 81%. The inhibition effect of pilicides is concentration dependent, which is a crucial property for their use as potential anti-bacterial agents. The data presented in this article indicate that pilicides in mM concentration effectively inhibit the adherence of Dr+ bacteria to the host cells, – the crucial, initial step in bacterial pathogenesis. Conclusions Structural analysis of the DraB chaperone clearly showed it to be a model of the FGL subfamily of chaperones. This permits us to conclude that analyzed pilicides in mM concentration are effective inhibitors of the assembly of adhesins belonging to the Dr family, and more speculatively, of other FGL-type adhesive organelles. The presented data and those published so far permit to speculate that based on the conservation of chaperone-usher pathway in Gram-negative bacteria , the pilicides are potential anti-bacterial agents with activity against numerous pathogens, the virulence of which is dependent on the adhesive structures of the chaperone-usher type.
    BMC Microbiology 06/2013; 13(1):131. DOI:10.1186/1471-2180-13-131 · 2.73 Impact Factor
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    • "The average all atom RMSD between the predicted and the native structures was about 7 ˚ A for the transmembrane domains of the β-barrel membrane proteins. However, it is unable to model structures when no homologous structural template is known or when a protein adopts a novel topology, such as VDAC (19 transmembrane β-strands) [30] and PapC (24 transmembrane β-strands) [31] proteins. A recent study described a method than can model the transmembrane domains with an average RMSD of 4 ˚ A for main Figure 1: Predicted structures of the transmembrane domains (in green) superimposed on experimentally determined structures (in red): (a) OmpF (pdb id:2omf), (b) OmpA (pdb id:1bxw), (c) BtuB (pdb id:1nqe), and (d) VDAC1 (pdb id:3emn). "
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    ABSTRACT: We briefly discuss recent progress in computational characterization of the sequence and structural properties of β-barrel membrane properties. We discuss the emerging concept of weakly stable regions in β-barrel membrane proteins, computational methods to identify these regions and mechanisms adopted by β-barrel membrane proteins in nature to stabilize them. We further discuss computational methods to identify protein-protein interactions in β-barrel membrane proteins and recent experimental studies that aim at altering the biophysical properties including oligomerization state and stability of β-barrel membrane proteins based on the emerging organization principles of these proteins from recent computational studies.
    Current pharmaceutical design 05/2013; 20(8). DOI:10.2174/13816128113199990071 · 3.45 Impact Factor
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