DNA packaging and ejection forces in bacteriophage

Abstract

We calculate the forces required to package (or, equivalently, acting to eject) DNA into (from) a bacteriophage capsid, as a function of the loaded (ejected) length, under conditions for which the DNA is either self-repelling or self-attracting. Through computer simulation and analytical theory, we find the loading force to increase more than 10-fold (to tens of piconewtons) during the final third of the loading process; correspondingly, the internal pressure drops 10-fold to a few atmospheres (matching the osmotic pressure in the cell) upon ejection of just a small fraction of the phage genome. We also determine an evolution of the arrangement of packaged DNA from toroidal to spool-like structures.T he classic Hershey-Chase experiment (1) of almost 50 years ago is best known for confirming DNA as the genetic material. But it was significant also as the demonstration that a bacterial virus (phage) leaves its protein capsid outside the cell it infects. More explicitly, upon binding to its receptor protein in the outer membrane of the bacterial cell, the viral capsid is opened and its DNA is injected into the cytoplasm. Obviously, this transfer can only happen as a passive process if the DNA is sufficiently pressurized in the capsid. For the past several decades, a great deal of experimental work has been devoted to determining the arrangement of ''packaged'' DNA in phage capsids through techniques that include x-ray scattering (2, 3), Raman spectroscopy (4), chemical cross-linking (5, 6), and electron microscopy (7,8,9). Various competing models have been proposed in which the DNA molecule is organized in concentric rings as a ''spool'' (3), in parallel segments joined at sharp kinks (10), or as a folded toroid (11). The most recent electron microscopy results on bacteriophages T7 (8) and T4 (9) show concentric ring structures that lend support to a spool-like structural motif. The underlying theoretical problem is also formidable, because one is confronted with the statistical-mechanical challenge of accounting for how a semif lexible, highly charged chain can be confined in dimensions comparable to its persistence length and yet hundreds of times smaller than its overall (contour) length L (12, 13). Although some estimates have been made of the pressure and elastic stress in a fully loaded capsid (12, 14), we are not aware of any attempt to treat the driving pressures during the course of ejection or the loading forces as a function of the extent of packaging (15). ¶In the present work, we connect the processes of loading DNA into, and ejecting it from, a phage capsid by calculating the energy of the chain as a function of the degree of loading or ejection. As already mentioned, the initial ejection is a passive process, being driven directly by the pressure difference inside and outside the capsid (16).ʈ Indeed, we shall show that the energy decreases monotonically as successively shorter lengths of DNA are left confined in the capsid; this rate of decrease corresponds to a force driving the chain outside. By ...

Biological consequences of tightly bent DNA: The other life of a macromolecular celebrity

“…This effect was taken into account in subsequent models of DNA packing. [16][17][18][28][29][30] These models are generally consistent with each other, but they focus on different kinds of predictions, such as the structure of the DNA, the forces and pressures involved in DNA confinement, and the effect of . This view is looking into the capsid at the portal (the entry site for DNA) and the green hoops reflect density corresponding to the packed DNA.…”

“…[14][15][16][17][18][19][20] The force due to bending is small during the initial stages of packing. However, as more DNA is forced into the capsid, the DNA takes up increasing amounts of available space and loops must be produced at smaller radii, increasing the force.…”

Free-living pathogens: Life-history constraints and strain competition

“…Charge repulsion between neighboring nucleic-acid segments stresses the capsid's integrity; a particle decays when internal forces, interacting with the physical environment, destabilize the capsid (Kindt et al, 2001). Smaller genomes (deletion mutants) decrease the internal force, and so increase virion persistence (Rubenstein, 1968;DePaepe and Taddei, 2006).…”

The conformation of double-stranded DNA inside bacteriophages depends on capsid size and shape

“…Some of these have used a statistical-thermodynamic approach to describe DNA packaging (Kindt et al, 2001;Purohit et al, 2005). Such models are capable of reproducing and predicting some thermodynamic properties such as packaging force, internal pressure, and internal energy.…”

“…Another group of studies overcomes this difficulty by simulating the packaging process, generating DNA conformations using coarsegrained DNA models and refining these by various molecular mechanics algorithms. These include molecular dynamics (MD) (Arsuaga et al, 2002;LaMarque et al, 2004;Locker, 2006;, Brownian dynamics (Kindt et al, 2001;Spakowitz and Wang, 2005) and Langevin dynamics (Forrey, 2006). Most of these studies have been limited to isometric (spherical or icosahedral) capsids and simple elastic DNA models, which precludes accurate prediction of thermodynamic properties.…”