Association Kinetics from Single Molecule Force Spectroscopy Measurements

Single molecule force spectroscopy is often used to study the dissociation of single molecules by applying mechanical force to the intermolecular bond. These measurements provide the kinetic parameters of dissociation. We present what to our knowledge is a new atomic force microscopy-based approach...

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Bibliographic Details
Published in:Biophysical journal 2009-04, Vol.96 (8), p.3412-3422
Main Authors: Guo, Senli, Lad, Nimit, Ray, Chad, Akhremitchev, Boris B.
Format: Article
Language:English
Subjects:
Algorithms
Binding sites
Biotin - metabolism
Data analysis
Kinetics
Measurement
Microscopy, Atomic Force
Models, Molecular
Molecules
Motion
Normal Distribution
Probability
Protein Binding
Spectroscopy, Imaging, and Other Techniques
Spectrum Analysis
Streptavidin - metabolism
Thermodynamics
Uncertainty
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Summary:Single molecule force spectroscopy is often used to study the dissociation of single molecules by applying mechanical force to the intermolecular bond. These measurements provide the kinetic parameters of dissociation. We present what to our knowledge is a new atomic force microscopy-based approach to obtain the activation energy of the association reaction and approximate grafting density of reactive receptors using the dependence of the probability to form molecular bonds on probe velocity when one of the interacting molecules is tethered by a flexible polymeric linker to the atomic force microscopy probe. Possible errors in the activation energy measured with this approach are considered and resulting corrections are included in the data analysis. This new approach uses the same experimental setup as traditional force spectroscopy measurements that quantify dissociation kinetics. We apply the developed methodology to measure the activation energy of biotin-streptavidin association (including a contribution from the steric factor) and obtain a value of 8 ± 1 kT. This value is consistent with the association rate measured previously in solution. Comparison with the solution-derived activation energy indicates that kinetics of biotin-streptavidin binding is mainly controlled by the reaction step.
ISSN:0006-3495
1542-0086
DOI:10.1016/j.bpj.2009.01.031