Biophysical Society Thematic Meeting - October 13-15, 2015

Biophysics of Proteins at Surfaces: Assembly, Activation, Signaling

Poster Abstracts

34-POS Board 34 Insights into HER2 Biology Through Structure-based Protein Engineering Jakob C. Stüber , Christian Jost, Iwo König, Yann Waltenspühl, Annemarie Honegger,

Benjamin Schuler, Andreas Plückthun. University of Zurich, Zürich, Switzerland.

Biparatopic Designed Ankyrin Repeat Proteins (DARPins) binding the oncoprotein HER2 outperform clinically approved antibody drugs in tissue culture experiments by a novel, apoptosis-inducing mechanism of cytotoxicity. X-ray structures of the DARPin moieties in complex with single extracellular subdomains require us to postulate previously unreported HER2 orientations to be induced by DARPin binding. Our work aims at elucidation of the molecular mechanism of biparatopic DARPins by three approaches: (1) Specific receptor labeling on live cells for fluorescence-based methods such as superresolution microscopy and fluorescence cross-correlation spectroscopy for dynamic and structural information; (2) overexpression and purification of HER2 from insect cells for structural investigations by electron microscopy; and (3) protein engineering to deliberately alter the DARPin properties and limit degrees of freedom within the DARPin-receptor complexes. Our current data are indeed in agreement with the adoption of non-native HER2 conformations upon DARPin binding, and furthermore suggest the formation of higher-order HER2 oligomers. These results might contribute to the formulation of a new therapeutic principle in receptor tyrosine kinase targeting.

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