Article

Dynamics of DNA Ejection from Bacteriophage

Division of Engineering and Applied Science, California Institute of Technology, Pasadena, California, USA.
Biophysical Journal (Impact Factor: 3.97). 08/2006; 91(2):411-20. DOI: 10.1529/biophysj.105.070532
Source: PubMed

ABSTRACT The ejection of DNA from a bacterial virus (i.e., phage) into its host cell is a biologically important example of the translocation of a macromolecular chain along its length through a membrane. The simplest mechanism for this motion is diffusion, but in the case of phage ejection a significant driving force derives from the high degree of stress to which the DNA is subjected in the viral capsid. The translocation is further sped up by the ratcheting and entropic forces associated with proteins that bind to the viral DNA in the host cell cytoplasm. We formulate a generalized diffusion equation that includes these various pushing and pulling effects and make estimates of the corresponding speedups in the overall translocation process. Stress in the capsid is the dominant factor throughout early ejection, with the pull due to binding particles taking over at later stages. Confinement effects are also investigated, in the case where the phage injects its DNA into a volume comparable to the capsid size. Our results suggest a series of in vitro experiments involving the ejection of DNA into vesicles filled with varying amounts of binding proteins from phage whose state of stress is controlled by ambient salt conditions or by tuning genome length.

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    • "The reasoning is the following. (1) DNA injection, which involves several biochemical events that may occur in parallel (Inamdar et al., 2006; Grayson and Molineux, 2007; Petrov et al., 2013), occurs in an environment that has sufficient variability to make responding to the environment a selective advantage via either (1) activation of the injection pathway that is most likely to succeed, or (2) reversal of injection when the environment is not favorable enough. Programmed, periodic stalling of injection provides time for feedback control. "
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    ABSTRACT: DNA packaging of phages phi29, T3 and T7 sometimes produces incompletely packaged DNA with quantized lengths, based on gel electrophoretic band formation. We discover here a packaging ATPase-free, in vitro model for packaged DNA length quantization. We use directed evolution to isolate a five-site T3 point mutant that hyper-produces tail-free capsids with mature DNA (heads). Three tail gene mutations, but no head gene mutations, are present. A variable-length DNA segment leaks from some mutant heads, based on DNase I-protection assay and electron microscopy. The protected DNA segment has quantized lengths, based on restriction endonuclease analysis: six sharp bands of DNA missing 3.7-12.3% of the last end packaged. Native gel electrophoresis confirms quantized DNA expulsion and, after removal of external DNA, provides evidence that capsid radius is the quantization-ruler. Capsid-based DNA length quantization possibly evolved via selection for stalling that provides time for feedback control during DNA packaging and injection.
    Virology 05/2014; s 456–457:157–170. DOI:10.1016/j.virol.2014.03.016 · 3.28 Impact Factor
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    • "The complementary ejection of the DNA into the host cell through small pores in the capsid is driven by the osmotic pressure exerted by the capsid and the internal stresses that have been built up in the highly-confined DNA polymer chain (Inamdar et al., 2006; Gelbart and Knobler, 2009). Because of its central importance in the life cycle of viruses, DNA ejection from a capsid has been intensively studied (Inamdar et al., 2006). Most theoretical treatments have investigated the role of microscopic mechanisms that arise naturally from the combined effects of osmotic pressure and bending energy. "
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    ABSTRACT: We construct a tractable model to describe the rate at which a knotted polymer is ejected from a spherical capsid via a small pore. Knots are too large to fit through the pore and must reptate to the end of the polymer for ejection to occur. The reptation of knots is described by symmetric exclusion on the line, with the internal capsid pressure represented by an additional biased particle that drives knots to the end of the chain. We compute the exact ejection speed for a finite number of knots L and find that it scales as 1/L. We establish a mapping to the solvable zero-range process. We also construct a continuum theory for many knots that matches the exact discrete theory for large L.
    Journal of Theoretical Biology 09/2009; 261(3):488-93. DOI:10.1016/j.jtbi.2009.08.021 · 2.30 Impact Factor
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    • "Single molecule experiments [25] have shown the pressure on the capsid walls to be on the order of 60 atm, inducing a significant resistance to the DNA encapsulation. These observations have generated a number of theoretical studies [26], primarily interested in the packaged structure [27] [28] [29] [30] [31], inter-strand spacing [32] [33] [34] [35], energy or pressure [36] [32] [27] [30] [37] [38] [39] [40], and the loading or ejection process [33] [41] [27] [28] [42] [30]. While the specific geometry of the confining viral capsid varies from phage to phage, the properties of the encapsulated DNA can be studied using spherical [29] [42] [28] or cylindrical [32] [34] confinement to a very good approximation. "
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    ABSTRACT: We develop an analytical method for studying the properties of a noninteracting wormlike chain (WLC) in confined geometries. The mean-field-like theory replaces the rigid constraints of confinement with average constraints, thus allowing us to develop a tractable method for treating a WLC wrapped on the surface of a sphere, and fully encapsulated within it. The efficacy of the theory is established by reproducing the exact correlation functions for a WLC confined to the surface of a sphere. In addition, the coefficients in the free energy are exactly calculated. We also describe the behavior of a surface-confined chain under external tension that is relevant for single molecule experiments on histone-DNA complexes. The force-extension curves display spatial oscillations, and the extension of the chain, whose maximum value is bounded by the sphere diameter, scales as f(-1) at large forces, in contrast to the unconfined chain that approaches the contour length as f(-1/2). A WLC encapsulated in a sphere, that is relevant for the study of the viral encapsulation of DNA, can also be treated using the mean-field approach. The predictions of the theory for various correlation functions are in excellent agreement with Langevin simulations. We find that strongly confined chains are highly structured by examining the correlations using a local winding axis. The predicted pressure of the system is in excellent agreement with simulations but, as is known, is significantly lower than the pressures seen for DNA packaged in viral capsids.
    Physical Review E 02/2009; 79(1 Pt 1):011924. DOI:10.1103/PhysRevE.79.011924 · 2.33 Impact Factor
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