Determination of the Kinetic On- and Off-Rate of Single Virus–Cell Interactions

Rankl, Christian; Wildling, Linda; Neundlinger, Isabel; Kienberger, Ferry; Gruber, Hermann J.; Blaas, Dieter; Hinterdorfer, Peter

doi:10.1007/978-1-61779-105-5_13

Cited by 19 publications

(16 citation statements)

References 27 publications

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“…For kinetic on-rate analysis, the BP (fraction of curves showing binding events) was determined at a certain hold time (t) (the time the tip is in contact with the surface). Those data were fitted and K D calculated as described previously 32 . In brief, the relationship between interaction time ( τ ) and BP is described by the following equation: where A is the maximum BP and t 0 the lag time.…”

Section: Methodsmentioning

confidence: 99%

Molecular interaction and inhibition of SARS-CoV-2 binding to the ACE2 receptor

et al. 2020

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Study of the interactions established between the viral glycoproteins and their host receptors is of critical importance for a better understanding of virus entry into cells. The novel coronavirus SARS-CoV-2 entry into host cells is mediated by its spike glycoprotein (S-glycoprotein), and the angiotensin-converting enzyme 2 (ACE2) has been identified as a cellular receptor. Here, we use atomic force microscopy to investigate the mechanisms by which the S-glycoprotein binds to the ACE2 receptor. We demonstrate, both on model surfaces and on living cells, that the receptor binding domain (RBD) serves as the binding interface within the S-glycoprotein with the ACE2 receptor and extract the kinetic and thermodynamic properties of this binding pocket. Altogether, these results provide a picture of the established interaction on living cells. Finally, we test several binding inhibitor peptides targeting the virus early attachment stages, offering new perspectives in the treatment of the SARS-CoV-2 infection.

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Section: Methodsmentioning

confidence: 99%

Molecular interaction and inhibition of SARS-CoV-2 binding to the ACE2 receptor

et al. 2020

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“…The kinetics for a receptor-ligand interaction (i.e., bonding and de-bonding) is characterized by the interaction’s on- and off-rates. We have estimated on-rates (k on ) of TIM-1−EBOV GP/VSV and TIM-4−EBOV GP/VSV interactions, using a method established by the Hinterdorfer group 23,48 . By assuming the interactions follow a pseudo first-order kinetics, the on-rates can be estimated using k on = (τ C eff ) −1 , where τ refers to the interaction time, and C eff denotes the effective concentration of the binding partner (in this case TIM-1 or TIM-4 on the surface) within an effective volume, V eff .…”

Section: Resultsmentioning

confidence: 99%

“…By assuming the interactions follow a pseudo first-order kinetics, the on-rates can be estimated using k on = (τ C eff ) −1 , where τ refers to the interaction time, and C eff denotes the effective concentration of the binding partner (in this case TIM-1 or TIM-4 on the surface) within an effective volume, V eff . The V eff can be approximated by the volume of a sphere, whose radius r eff is the sum of the size of the virus and cross-linker 48 . Therefore, the equation can be rewritten as: k on = N A ·(4/3)·π·r eff 3·n b −1 ·τ −1 , where N A is the Avogadro constant, and n b is the number of the binding partner 48 .…”

Section: Resultsmentioning

confidence: 99%

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Biomechanical characterization of TIM protein–mediated Ebola virus–host cell adhesion

Dragovich

Fortoul

Jagota

et al. 2019

Sci Rep

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Since the most recent outbreak, the Ebola virus (EBOV) epidemic remains one of the world’s public health and safety concerns. EBOV is a negative-sense RNA virus that can infect humans and non-human primates, and causes hemorrhagic fever. It has been proposed that the T-cell immunoglobulin and mucin domain (TIM) family proteins act as cell surface receptors for EBOV, and that the interaction between TIM and phosphatidylserine (PS) on the surface of EBOV mediates the EBOV–host cell attachment. Despite these initial findings, the biophysical properties of the TIM-EBOV interaction, such as the mechanical strength of the TIM-PS bond that allows the virus-cell interaction to resist external mechanical perturbations, have not yet been characterized. This study utilizes single-molecule force spectroscopy to quantify the specific interaction forces between TIM-1 or TIM-4 and the following binding partners: PS, EBOV virus-like particle, and EBOV glycoprotein/vesicular stomatitis virus pseudovirion. Depending on the loading rates, the unbinding forces between TIM and ligands ranged from 40 to 100 pN, suggesting that TIM-EBOV interactions are mechanically comparable to previously reported adhesion molecule–ligand interactions. The TIM-4–PS interaction is more resistant to mechanical force than the TIM-1–PS interaction. We have developed a simple model for virus–host cell interaction that is driven by its adhesion to cell surface receptors and resisted by membrane bending (or tension). Our model identifies critical dimensionless parameters representing the ratio of deformation and adhesion energies, showing how single-molecule adhesion measurements relate quantitatively to the mechanics of virus adhesion to the cell.

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“…Longer contact times of the MFZ-AFM tip on the cellular surface resulted in a significant increase in the binding probability. Assuming that the ligand-transporter interaction can be approximated with first-order kinetics, an estimation of the kinetic on-rate constant k on from single molecule unbinding force measurements requires that the interaction time τ and the effective concentration c eff be determined for k on = (τ × c eff ) −1 (29, 32, 46). The interaction time was calculated from the binding probability at different dwell times by using p = A (1 − exp(−( t − t 0 )/τ)), with t 0 representing the lag time and with A representing the maximum achievable binding probability.…”

Section: Resultsmentioning

confidence: 99%

Probing Binding Pocket of Serotonin Transporter by Single Molecular Force Spectroscopy on Living Cells

Wildling

Rankl

Haselgrübler³

et al. 2012

Journal of Biological Chemistry

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Background: The serotonin transporter is the site of action of antidepressants and amphetamines.Results: Single molecular force spectroscopy allowed for mapping the energy landscape involved in MFZ2-12/SERT binding.Conclusion: Our data indicate that the outer vestibule imposes a barrier on the entry of MFZ2-12 into the SERT substrate binding site.Significance: Our results provide a useful framework for a further exploration of antidepressant binding.

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Determination of the Kinetic On- and Off-Rate of Single Virus–Cell Interactions

Cited by 19 publications

References 27 publications

Molecular interaction and inhibition of SARS-CoV-2 binding to the ACE2 receptor

Molecular interaction and inhibition of SARS-CoV-2 binding to the ACE2 receptor

Biomechanical characterization of TIM protein–mediated Ebola virus–host cell adhesion

Probing Binding Pocket of Serotonin Transporter by Single Molecular Force Spectroscopy on Living Cells

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