Direct measurement of the interaction force between immunostimulatory CpG-DNA and TLR9 fusion protein

The specific interaction between human Toll‐like receptor 9 (TLR9)–ectodomain (ECD)–fusion protein and immunostimulatory CpG–DNA was measured using force spectroscopy. Flexible tethers were used between receptors and surface as well as between DNA and atomic force microscope tip to make efficient re...

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Bibliographic Details
Published in:Journal of molecular recognition 2012-02, Vol.25 (2), p.74-81
Main Authors: Klein, Dionne C. G., Øvrebø, Kirsti Marie, Latz, Eicke, Espevik, Terje, Stokke, Bjørn T.
Format: Article
Language:English
Subjects:
DNA - metabolism
force spectroscopy
Humans
Immunization
innate immune receptor binding
kinetics
Microscopy, Atomic Force - methods
Models, Biological
Oligodeoxyribonucleotides - metabolism
Protein Binding
Recombinant Fusion Proteins - metabolism
single molecular pair interaction
Spectrum Analysis
Thermodynamics
Toll-like receptor 9
Toll-Like Receptor 9 - metabolism
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Summary:The specific interaction between human Toll‐like receptor 9 (TLR9)–ectodomain (ECD)–fusion protein and immunostimulatory CpG–DNA was measured using force spectroscopy. Flexible tethers were used between receptors and surface as well as between DNA and atomic force microscope tip to make efficient recognition studies possible. The molecular recognition forces detected are in the range of 50 to 150 ± 20 pN at the used force‐loading rates, and the molecular interaction probability was much reduced when the receptors were blocked with free CpG–DNA. A linear increase of the unbinding force with the logarithm of the loading rate was found over the range 0.1 to 30 nN/s. This indicates a single potential barrier characterizing the energy landscape and no intermediate state for the unbinding pathway of CpG–DNA from the TLR9–ECD. Two important kinetic parameters for CpG–DNA interaction with TLR9–ECD were determined from the force‐loading rate dependency: an off‐rate of koff = 0.14 ± 0.10 s‐1 and a binding interaction length of xβ = 0.30 ± 0.03 nm, which are consistent with literature values. Various models for the molecular interaction of this innate immune receptor binding to CpG–DNA are discussed. Copyright © 2012 John Wiley & Sons, Ltd.
ISSN:0952-3499
1099-1352
DOI:10.1002/jmr.2156