Live cell measurements of interaction forces and binding kinetics between Discoidin Domain Receptor 1 (DDR1) and collagen I with atomic force microscopy.

Autor: Sarkar A; Department of Physics and Astronomy, Wayne State University, Detroit, MI, United States., Sohail A; Department of Pathology and Oncology, Wayne State University, Detroit, MI, United States., Dong J; Department of Physics and Astronomy, Wayne State University, Detroit, MI, United States., Prunotto M; School of Pharmaceutical Sciences, University of Geneva, Switzerland., Shinki K; Department of Mathematics, Wayne State University, Detroit, MI, United States., Fridman R; Department of Pathology and Oncology, Wayne State University, Detroit, MI, United States., Hoffmann PM; Department of Physics and Astronomy, Wayne State University, Detroit, MI, United States. Electronic address: hoffmann@wayne.edu.
Jazyk: angličtina
Zdroj: Biochimica et biophysica acta. General subjects [Biochim Biophys Acta Gen Subj] 2019 Nov; Vol. 1863 (11), pp. 129402. Date of Electronic Publication: 2019 Jul 24.
DOI: 10.1016/j.bbagen.2019.07.011
Abstrakt: Background: Discoidin Domain Receptors (DDRs) are membrane-tethered proteins of the receptor tyrosine kinase family, which signal in response to collagen. DDR expression is associated with human diseases, including fibrosis and cancer. The role of DDRs in human pathogenesis is mediated by dysregulated receptor function in response to the collagenous milieu. Thus, understanding DDR-collagen interactions is important for developing novel therapeutic strategies against DDRs.
Methods: We developed a biophysical method to isolate and measure specific interactions between DDR1 and collagen in live cells at the single molecule level using atomic force microscopy. This new method is capable of providing density and kinetics of membrane receptors in live cells.
Results: We isolated DDR1-collagen interactions and quantified the association and dissociation rates of the DDR1-collagen I complex. We estimated separate binding probabilities of collagen I to DDR and integrin, and by combining kinetic and binding probability data, we were able to estimate the density of receptors in two cancer cell types. We also tested the viability of a DDR1 blocking antibody and determined its efficacy in suppressing DDR1-collagen binding.
Conclusions: The new method shows promise in quantifying receptor-ligand kinetics and receptor density on live cells.
General Significance: The new approach is applicable to other receptor-ligand systems and allows the determination of critical parameters at the single cell/single molecule level - in particular, the direct determination of kinetic and density differences of receptors in different cell types. This capability should prove to be useful in cancer research and drug design.
(Copyright © 2019 Elsevier B.V. All rights reserved.)
Databáze: MEDLINE
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