TY - JOUR
T1 - Quantifying the Interaction between EGFR Dimers and Grb2 in Live Cells
AU - Del Piccolo, Nuala
AU - Hristova, Kalina
N1 - Publisher Copyright:
© 2017 Biophysical Society
PY - 2017/9/19
Y1 - 2017/9/19
N2 - Adaptor proteins are a class of cytoplasmic proteins that bind to phosphorylated residues in receptor tyrosine kinases and trigger signaling cascades that control critically important cellular processes, such as cell survival, growth, differentiation, and motility. Here, we seek to characterize the interaction between epidermal growth factor receptor (EGFR) and the cytoplasmic adaptor protein growth factor receptor-bound protein 2 (Grb2) in a cellular context. To do so, we explore the utility of a highly biologically relevant model system, mammalian cells under reversible osmotic stress, and a recently introduced Förster resonance energy transfer microscopy method, fully quantified spectral imaging. We present a method that allows us to quantify the stoichiometry and the association constant of the EGFR-Grb2 binding interaction in the plasma membrane, in the presence and absence of activating ligand. The method that we introduce can have broad utility in membrane protein research, as it can be applied to different membrane protein-cytoplasmic protein pairs.
AB - Adaptor proteins are a class of cytoplasmic proteins that bind to phosphorylated residues in receptor tyrosine kinases and trigger signaling cascades that control critically important cellular processes, such as cell survival, growth, differentiation, and motility. Here, we seek to characterize the interaction between epidermal growth factor receptor (EGFR) and the cytoplasmic adaptor protein growth factor receptor-bound protein 2 (Grb2) in a cellular context. To do so, we explore the utility of a highly biologically relevant model system, mammalian cells under reversible osmotic stress, and a recently introduced Förster resonance energy transfer microscopy method, fully quantified spectral imaging. We present a method that allows us to quantify the stoichiometry and the association constant of the EGFR-Grb2 binding interaction in the plasma membrane, in the presence and absence of activating ligand. The method that we introduce can have broad utility in membrane protein research, as it can be applied to different membrane protein-cytoplasmic protein pairs.
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U2 - 10.1016/j.bpj.2017.06.029
DO - 10.1016/j.bpj.2017.06.029
M3 - Article
C2 - 28734476
AN - SCOPUS:85024841801
SN - 0006-3495
VL - 113
SP - 1353
EP - 1364
JO - Biophysical journal
JF - Biophysical journal
IS - 6
ER -