DNA Dendron Tagging\nas a Universal Way to Deliver\nProteins to Cells

The use of proteins as intracellular probes and therapeutic\ntools\nis often limited by poor intracellular delivery. One approach to enabling\nintracellular protein delivery is to transform proteins into spherical\nnucleic acid (proSNA) nanoconstructs, with surfaces chemically modified\nwith a dense shell of radially oriented DNA that can engage with cell-surface\nreceptors that facilitate endocytosis. However, proteins often have\na limited number of available reactive surface residues for DNA conjugation\nsuch that the extent of DNA loading and cellular uptake is restricted.\nIndeed, DNA surface density and sequence have been correlated with\nscavenger-receptor engagement, the first step of cellular internalization.\nHere, we report how branched DNA dendrons with dibenzocyclooctyne\ngroups and proteins genetically engineered to include the noncanonical\namino acid azido-phenylalanine for click chemistry can be used to\nsynthesize hybrid DNA dendron-protein architectures that exhibit outstanding\ncellular internalization properties, without the need for extensive\nsurface modification. In a head-to-head comparison, protein–DNA\ndendron structures (where DNA is concentrated in a local area) are\ntaken up by cells more rapidly and to a greater extent than proSNAs\n(where the DNA is evenly distributed). Also, protein-G-rich dendron\nstructures show enhanced uptake compared to protein-T-rich dendron\nstructures, highlighting the importance of oligonucleotide sequence\non nanoconjugate uptake. Finally, a generalizable method for chemically\ntagging proteins with dendrons that does not require mutagenesis is\ndescribed. When a range of proteins, spanning 42 to 464 kDa, were\nmodified through surface lysines with this method, a significant increase\nin their cellular uptake (up to 17-fold) compared to proteins that\nare not coupled to a DNA dendron was observed.