DNA confined by surfaces

University Of Texas Med Br Galveston, Galveston TX

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Abstract

DESCRIPTION (provided by applicant): Better understanding the biophysical basis of the biological process to transfer a viral genome to infect a cell is important to many disease related fields. Predicting the thermodynamic pressures and forces including the osmotic pressure necessary to confine DNA-a highly-negatively charged, elastic polymer-into capsids (over a 250-fold compaction) is a problem with implications not only relevant to infectious disease mechanism but in phage therapy or phage antibiotics(1) and therapeutic delivery(2). Experimental measurements of phage DNA confinement include osmotic pressure ejection-inhibition experiments(3) and single-molecule loading force measurements that provide force or pressure data for validation of theoretical models.(4, 5) Structural insight into DNA packaging is aided by cryo-electron microscopy asymmetric reconstructions done in the NCMI with our collaborator Chiu.(6, 7) Most current models of phage packing assume DNA behaves as a linearly elastic polymer that bends uniformly under stress, like the 'inverse spool' model.(8) The assumption of such spooled conformations is based primarily on interpretations of cryo-EM density maps, obtained by averaging thousands of structures(9), which show density rings, especially near the capsid surface. Recent evidence shows that during translocation packing, the DNA helix is rotated in a left-handed direction thus under twisting it.(10, 11) It Is known tha under twisting reduces persistence length by 2 orders of magnitude when strand separation occurs in sequence specific places.(12) Our hypothesis is that DNA kinking induced disorder can have a strong effect on packing and pressures. How DNA overcomes the unfavorable thermodynamic barrier to enter and pack inside a capsid depends on many different intermolecular interactions. Because phage genomes are around ten kilo-basepairs long, we will employ a multi scale technique to model the structure and consequent thermodynamics. We will refine a coarse-grained model of DNA from our previous work.(13) Preliminary simulations of unconnected DNA coarse grained polymer beads in capsid-like confinement already show ringed density distributions consistent with cryo-EM data. Connected polymer paths will be constructed consistent with data. We will produce an ensemble of entropically-driven, low free energy conformations of DNA in confinement. Ultimately, we will test hypotheses related to the amount of disorder, ion screening and the contribution of DNA-protein confinement interactions.