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Beutel, O.* ; Roder, F.* ; Birkholz, O.* ; Rickert, C.* ; Steinhoff, H.* ; Grzybek, M. ; Coskun, Ü. ; Piehler, J.*

Two-dimensional trap for ultrasensitive quantification of transient protein interactions.

ACS Nano 9, 9783-9791 (2015)
Verlagsversion DOI
Open Access Green möglich sobald Postprint bei der ZB eingereicht worden ist.
We present an ultrasensitive technique for quantitative protein–protein interaction analysis in a two-dimensional format based on phase-separated, micropatterned membranes. Interactions between proteins captured to lipid probes via an affinity tag trigger partitioning into the liquid-ordered phase, which is readily quantified by fluorescence imaging. Based on a calibration with well-defined low-affinity protein–protein interactions, equilibrium dissociation constants >1 mM were quantified. Direct capturing of proteins from mammalian cell lysates enabled us to detect homo- and heterodimerization of signal transducer and activator of transcription proteins. Using the epidermal growth factor receptor (EGFR) as a model system, quantification of low-affinity interactions between different receptor domains contributing to EGFR dimerization was achieved. By exploitation of specific features of the membrane-based assay, the regulation of EGFR dimerization by lipids was demonstrated.
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Publikationstyp Artikel: Journalartikel
Dokumenttyp Wissenschaftlicher Artikel
Schlagwörter Protein-protein Interaction ; Polymer-supported Membrane ; Lipid Phase Separation ; Fluorescence Microscopy ; Signaling Complexes ; Protein-lipid Interaction
ISSN (print) / ISBN 1936-0851
e-ISSN 1936-086X
Zeitschrift ACS Nano
Quellenangaben Band: 9, Heft: 10, Seiten: 9783-9791 Artikelnummer: , Supplement: ,
Verlag American Chemical Society (ACS)
Begutachtungsstatus Peer reviewed
Institut(e) Institute for Pancreatic Beta Cell Research (IPI)