Tailoring\nDNA Origami Protection: A Study of Oligolysine-PEG\nCoatings for Improved Colloidal, Structural, and Functional Integrity

Application of protective polymer coatings to enhance\nthe biostability\nof DNA-based nanomaterials (DONs) has become common practice in <i>in vitro</i> and <i>in vivo</i> experiments. While\nthe functional effect of these coatings is obvious, a detailed molecular\npicture of what is protected and for how long remains unclear. Additionally,\nthe use of the oligolysine-1kPEG protective polymer has been limited\ndue to aggregation issues. In this study, we evaluated the colloidal\nstability, structural integrity, and functional preservation of DONs\ncoated with oligolysine (K)-1k/5kPEG block copolymers. Dynamic light\nscattering and transmission electron microscopy were employed to assess\ncolloidal stability before and after degradation. A FRET-based assay\nwas developed to monitor the directionality of degradation, while\nquantitative PCR measured the protection of functional DNA handles,\ncrucial for the design of ligand-functionalized DONs. Our results\nshow that K<sub>10</sub>-1kPEG, while prone to aggregation, can offer\nsimilar protection against nucleases as K<sub>10</sub>-5kPEG, provided\nbuffer conditions are carefully chosen. Maintaining the colloidal,\nstructural, and functional stability before and after nuclease exposure\nsupports DON applications, particularly at the biointerface. These\ninsights provide valuable guidelines for selecting the most effective\nprotection strategy and enhancing DON functionality across diverse\nbiological environments.