Retroviral integration into topologically-interlocked DNAs to probe the role of DNA structure and screen viral inhibitors

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Topology-specific DNA structures, such as small loops, supercoils, knots, and catenated mini and maxicircles, occur in numerous instances in vivo. Besides their presence in vivo, synthetic molecules with interlocked units were achieved in macromolecular chemistry decades ago and are the current topic of interest in structural DNA nanotechnology. Due to the circular nature of these minicircles and interlocked DNAs, their structure and topology resemble nucleosomal DNA. Therefore, DNA minicircles with and without interlocked structures are attractive targets to probe the topological features of DNA and DNA-protein interactions. In this study, we aimed to design and prepare the DNA minicircles in free and topologically interlocked forms. These minicircles mimic nucleosomal DNA in terms of size and structure. Also, we have designed a frame-shaped DNA origami nanostructure and incorporated the topologically interlocked rotaxanes and catenanes. During our study, we noticed that these minicircles/interlocked structures and also the DNA origami frame are relatively unstable under application-specific conditions. Thus, before the investigation of the DNA-protein interactions and drug screening, it was necessary to stabilize these structures. We spent a reasonable amount of time in developing the stabilization methods for these DNA structures, and finally achieved stable substrates for the DNA-protein interactions and drug screening.