DNA confined by surfaces

DNA 受表面限制

• 批准号: 8593300
• 负责人: Bernard MONTGOMERY PETTITT
• 金额: $ 26.78万
• 依托单位: UNIVERSITY OF TEXAS MED BR GALVESTON
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-04-01 至 2016-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8593300
• 关键词: Antibiotics; Bacteriophages; Biochemical Process; Biological Process; Capsid; Cells; Cereals; Charge; Communicable Diseases; Coupling; Cryoelectron Microscopy; DNA; DNA Packaging; Data; Disease; Elasticity; Electrostatics; Free Energy; Genome; Geometry; Goals; Grant; Hand; Hydrostatic Pressure; Infection; Integral Equation; Ions; Lead; Left; Length; Literature; Maps; Measurement; Measures; Methods; Modeling; Molecular; Molecular Conformation; Motor; Nature; Nucleic Acids; Osmotic Pressure; Polymers; Property; Proteins; Saline; Sampling; Series; Simulate; Solutions; Stress; Structure; Surface; System; Techniques; Testing; Theoretical model; Therapeutic; Thermodynamics; Validation; Viral Genome; Work; base; density; driving force; insight; intermolecular interaction; kinked DNA; multi-scale modeling; planetary Atmosphere; pressure; public health relevance; reconstruction; research study; screening; simulation; single molecule; success; theories

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.

描述（由申请人提供）：更好地了解转移病毒基因组以感染细胞的生物过程的生物物理基础对于许多相关疾病很重要 字段。预测热力学压力和力，包括将 DNA（一种带高负电荷的弹性聚合物）限制在衣壳中（压缩超过 250 倍）所需的渗透压，是一个不仅与传染病机制相关而且与噬菌体治疗相关的问题。或噬菌体抗生素(1) 和治疗递送(2)。噬菌体 DNA 限制的实验测量包括渗透压喷射抑制实验 (3) 和单分子加载力测量，为验证理论模型提供力或压力数据。(4, 5) 通过低温辅助了解 DNA 包装的结构与我们的合作者 Chiu 在 NCMI 进行的电子显微镜不对称重建。(6, 7) 当前大多数噬菌体堆积模型都假设 DNA 表现为可弯曲的线性弹性聚合物在应力下均匀地分布，如“逆线轴”模型。(8) 这种线轴构象的假设主要基于对冷冻电镜密度图的解释，该图是通过对数千个结构进行平均而获得的(9)，这些结构显示出密度环，尤其是在附近衣壳表面。最近的证据表明，在易位包装过程中，DNA 螺旋沿左手方向旋转，从而使其受到扭曲。(10, 11) 众所周知，当序列特异性发生链分离时，扭曲会使持久长度减少 2 个数量级(12) 我们的假设是 DNA 扭结引起的紊乱会对包装和压力产生强烈影响。 DNA 如何克服不利的热力学障碍进入并包装在衣壳内取决于许多不同的分子间相互作用。由于噬菌体基因组长约十公斤碱基对，因此我们将采用多尺度技术来模拟其结构和随后的热力学。我们将从我们之前的工作中完善一个粗粒度的 DNA 模型。(13) 对衣壳状限制中未连接的 DNA 粗粒度聚合物珠的初步模拟已经显示出与冷冻电镜数据一致的环形密度分布。连接的聚合物路径将根据数据构建。我们将在限制条件下产生一组由熵驱动、低自由能的 DNA 构象。最终，我们将测试与无序量、离子筛选和 DNA-蛋白质限制相互作用的贡献相关的假设。