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.
Understanding the biophysical basis of the biological process which transfers a viral genome to infect a cell is important to many disease related fields. Predicting the thermodynamic pressures including the osmotic pressure necessary to confine DNA in phage capsids (over a 250-fold compaction) is a problem with implications in infection, phage therapies and therapeutic delivery. We will resolve questions of the thermodynamic mechanism of DNA ejection by phages.
|Wang, Qian; Irobalieva, Rossitza N; Chiu, Wah et al. (2017) Influence of DNA sequence on the structure of minicircles under torsional stress. Nucleic Acids Res 45:7633-7642|
|Myers, Christopher G; Pettitt, B Montgomery (2017) Phage-like packing structures with mean field sequence dependence. J Comput Chem 38:1191-1197|
|Esadze, Alexandre; Chen, Chuanying; Zandarashvili, Levani et al. (2016) Changes in conformational dynamics of basic side chains upon protein-DNA association. Nucleic Acids Res 44:6961-70|
|Chen, Chuanying; Pettitt, B Montgomery (2016) DNA Shape versus Sequence Variations in the Protein Binding Process. Biophys J 110:534-544|
|Bates, David; Pettitt, B Montgomery; Buck, Gregory R et al. (2016) Importance of disentanglement and entanglement during DNA replication and segregation: Comment on: ""Disentangling DNA molecules"" by Alexander Vologodskii. Phys Life Rev 18:160-164|
|Wang, Qian; Pettitt, B Montgomery (2016) Sequence Affects the Cyclization of DNA Minicircles. J Phys Chem Lett 7:1042-6|
|Wang, Qian; Myers, Christopher G; Pettitt, B Montgomery (2015) Twist-induced defects of the P-SSP7 genome revealed by modeling the cryo-EM density. J Phys Chem B 119:4937-43|
|Wang, Qian; Pettitt, B Montgomery (2014) Modeling DNA thermodynamics under torsional stress. Biophys J 106:1182-93|
|Myers, Christopher G; Pettitt, B Montgomery (2013) Communication: Origin of the contributions to DNA structure in phages. J Chem Phys 138:071103|
|Theruvathu, Jacob A; Yin, Y Whitney; Pettitt, B Montgomery et al. (2013) Comparison of the structural and dynamic effects of 5-methylcytosine and 5-chlorocytosine in a CpG dinucleotide sequence. Biochemistry 52:8590-8|
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