Visualization and targeted disruption of protein interactions in living cells

Henry D. Herce, Wen Deng, Jonas Helma, Heinrich Leonhardt () and M. Cristina Cardoso ()
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Henry D. Herce: Technische Universität Darmstadt
Wen Deng: Center for Integrated Protein Science Munich, Ludwig Maximilians University Munich
Jonas Helma: Center for Integrated Protein Science Munich, Ludwig Maximilians University Munich
Heinrich Leonhardt: Center for Integrated Protein Science Munich, Ludwig Maximilians University Munich
M. Cristina Cardoso: Technische Universität Darmstadt

Nature Communications, 2013, vol. 4, issue 1, 1-8

Abstract: Abstract Protein–protein interactions are the basis of all processes in living cells, but most studies of these interactions rely on biochemical in vitro assays. Here we present a simple and versatile fluorescent-three-hybrid (F3H) strategy to visualize and target protein–protein interactions. A high-affinity nanobody anchors a GFP-fusion protein of interest at a defined cellular structure and the enrichment of red-labelled interacting proteins is measured at these sites. With this approach, we visualize the p53–HDM2 interaction in living cells and directly monitor the disruption of this interaction by Nutlin 3, a drug developed to boost p53 activity in cancer therapy. We further use this approach to develop a cell-permeable vector that releases a highly specific peptide disrupting the p53 and HDM2 interaction. The availability of multiple anchor sites and the simple optical readout of this nanobody-based capture assay enable systematic and versatile analyses of protein–protein interactions in practically any cell type and species.

Date: 2013
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DOI: 10.1038/ncomms3660

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