RT Journal Article SR Electronic T1 Cytosolic delivery of large supramolecular protein complexes arranged on DNA nanopegboards JF bioRxiv FD Cold Spring Harbor Laboratory SP 236729 DO 10.1101/236729 A1 Pavel M. Nikolov A1 Katja J. Koßmann A1 Alessa Schilling A1 Alessandro Angelin A1 Josipa Brglez A1 Alina Klein A1 Robert Tampé A1 Kersten S. Rabe A1 Christof M. Niemeyer YR 2017 UL http://biorxiv.org/content/early/2017/12/19/236729.abstract AB A generic methodology for cytosolic delivery of large supramolecular multiprotein complexes into living cells is described that takes advantage of the highly-controllable bottom-up fabrication of protein-decorated DNA nanostructures and the microfluidic “cell squeezing” technique. Therein, cells are deformed upon passage through a narrow constriction leading to formation of transient holes in the cell membrane that enable the diffusion of the protein-DNA nanostructures from the surrounding buffer into the cytosol. A diverse set of multiprotein complexes was assembled on DNA origami nanostructures using streptavidin and the sensitive glucose sensor protein FLIP as model systems. We demonstrate that our approach allows for the direct cytosolic delivery of these multifunctional protein complexes into the cytosol of HeLa cells. We also demonstrate that targeting groups can be incorporated into the protein-DNA nanoassemblies to enable their intracellular targeting to cytosolic compartments, such as the cytoskeleton or nucleus. We believe that this methodology will open up novel strategies for research in fundamental cell biology, such as the reverseengineering of the supramolecular machinery involved in gene regulation, cell signalling, or cell division. Furthermore, direct applications in immunotherapy can be foreseen.