Label-free measuring and mapping of binding kinetics of membrane proteins in single living cells

Abstract

Membrane proteins mediate a variety of cellular responses to extracellular signals. Although membrane proteins are studied intensively for their values as disease biomarkers and therapeutic targets, in situ investigation of the binding kinetics of membrane proteins with their ligands has been a challenge. Traditional approaches isolate membrane proteins and then study them ex situ, which does not reflect accurately their native structures and functions. We present a label-free plasmonic microscopy method to map the local binding kinetics of membrane proteins in their native environment. This analytical method can perform simultaneous plasmonic and fluorescence imaging, and thus make it possible to combine the strengths of both label-based and label-free techniques in one system. Using this method, we determined the distribution of membrane proteins on the surface of single cells and the local binding kinetic constants of different membrane proteins. Furthermore, we studied the polarization of the membrane proteins on the cell surface during chemotaxis.

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Figure 1: Surface plasmon resonance microscopy.
Figure 2: Glycoprotein–lectin interaction.
Figure 3: Mapping glycoprotein distribution and binding kinetics.
Figure 4: Glycoprotein polarization during chemotaxis.
Figure 5: Mapping nAChRs.

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Acknowledgements

We thank the National Institutes of Health (R21RR026235) for support.

Author information

W.W. and Y.Y. designed and performed the experiments. W.W., Y.Y., S.W., V.J.N. and N.J.T. discussed the results. Q.L. and J.W. provided the cell lines and helped with the immunofluorescence of nAChR. W.W. and N.J.T. wrote the paper. N.J.T. conceived the experiment and supervised the project.

Correspondence to Nongjian Tao.

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The authors declare no competing financial interests.

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Wang, W., Yang, Y., Wang, S. et al. Label-free measuring and mapping of binding kinetics of membrane proteins in single living cells. Nature Chem 4, 846–853 (2012). https://doi.org/10.1038/nchem.1434

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