DNA Dendron Tagging as a Universal Way to Deliver Proteins to Cells

The use of proteins as intracellular probes and therapeutic tools is often limited by poor intracellular delivery. One approach to enabling intracellular protein delivery is to transform proteins into spherical nucleic acid (proSNA) nanoconstructs, with surfaces chemically modified with a dense shell of radially oriented DNA that can engage with cell-surface receptors that facilitate endocytosis. However, proteins often have a limited number of available reactive surface residues for DNA conjugation such that the extent of DNA loading and cellular uptake is restricted. Indeed, DNA surface density and sequence have been correlated with scavenger-receptor engagement, the first step of cellular internalization. Here, we report how branched DNA dendrons with dibenzocyclooctyne groups and proteins genetically engineered to include the noncanonical amino acid azido-phenylalanine for click chemistry can be used to synthesize hybrid DNA dendron-protein architectures that exhibit outstanding cellular internalization properties, without the need for extensive surface modification. In a head-to-head comparison, protein–DNA dendron structures (where DNA is concentrated in a local area) are taken up by cells more rapidly and to a greater extent than proSNAs (where the DNA is evenly distributed). Also, protein-G-rich dendron structures show enhanced uptake compared to protein-T-rich dendron structures, highlighting the importance of oligonucleotide sequence on nanoconjugate uptake. Finally, a generalizable method for chemically tagging proteins with dendrons that does not require mutagenesis is described. When a range of proteins, spanning 42 to 464 kDa, were modified through surface lysines with this method, a significant increase in their cellular uptake (up to 17-fold) compared to proteins that are not coupled to a DNA dendron was observed.