Experimental and Theoretical Studies of DNA-Macroion Interactions.

[摘要] The thesis uses experiments and simulations to examine the interactions of DNA molecules with proteins and protein-like nanoparticles, with applications to protein search and targeting of DNA sequences, and to DNA complexation in chromatin and for gene delivery.Two topics are covered in depth.In the first topic, kinetic Monte Carlo simulations, one dimensional reaction-diffusion equations,and analytical methods are used to determine rate at which DNA-binding proteins (e.g. transcription factors) can find the target sequences in long DNA molecules through a combination of sequence-dependent 1D diffusion and sequence-independent 3D diffusion. We quantify how thousands of ``decoy sites;;;; which have similar base pair sequences as target sites slow down the protein targeting process dramatically. We find the conditions under which the protein targeting process can be sped-up, including the effect of a ``two-state;;;; protein model, allowing for both rapid diffusion and accurate searching. In the second topic, we investigate how the surface charge density of a poly(amido amine) (or PAMAM) dendrimer affects its ability to condense on DNA,using light scattering, circular dichroism,and single molecule imaging of dendrimer-DNA complexes combed onto surfaces and tethered to those surfaces under flow. This study is important not only for understandinghow to condense dsDNA to facilitate its penetration into cell membranes for non-viral gene therapy, but also because PAMAM dendrimers provide an ideal biomimic of DNA-binding proteins (e.g. histones). To describe DNA compaction by dendrimers, we develop a mesoscale model combining a coarse-grained DNA model of de Pablo and coworkers which resolves the DNA double helix structure with acoarse-grained dendrimer model of Muthukumar and coworkers. The predictions of our new model on effects of dendrimer generation, dendrimer surface charge density, and salt concentration on dendrimer-DNA complexes formation are consistent with both experiments and potential of mean force results from all-atom molecular dynamics simulations, but give much more detail regarding the structure of the complex. Moreover, this model can be extended to other cationic macroion-DNA systems which are also of great interest, such as, polylysine, micelles, and colloidal particles.

[发布机构] University of Michigan

[效力级别] PAMAM dendrimer

[关键词] DNA condensation;PAMAM dendrimer;Chemical Engineering;Engineering;Chemical Engineering