Insights into the Role of Nucleic Acid Structure and Topology in Controlling Condensation
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DNA condensation is a fundamental process in all living organisms. The highly abundant nucleoid-associated proteins, HU and IHF, present in bacteria, have been shown to play an important role in shaping the nucleoid. However, the exact mechanism is not well understood. In this thesis, we have demonstrated that both HU and IHF guide DNA to condense into linear bundle-like structures in presence of cellular condensing components, but the proteins alone do not condense DNA into densely packed structures. Our results suggest a mechanism by which HU and IHF could act as architectural proteins during in vitro and in vivo DNA condensation. More recently, DNA condensation has attracted much attention for its relevance in optimizing artificial DNA delivery systems for gene therapy. The research presented in this dissertation provides in depth biophysical studies that demonstrate how local modulations in the nucleic acid structure can be used to control both the size and the morphology DNA condensates. We describe a novel strategy for improving the condensation of oligonucleotides that is based on the self-organization of half-sliding complementary oligonucleotides into long duplexes (ca. kb) with flexible sites at regular intervals along the duplex backbones, in the form of single-stranded nicks or single-stranded gaps. Our results also provide new insights into the role of DNA flexibility in condensate formation and suggest the potential for the use of this DNA structure in the design of vectors for oligonucleotide and gene delivery.