Single-molecule experiments with optical tweezers have become an important tool to study the properties and mechanisms of biological systems, such as cells and nucleic acids. In particular, force unzipping experiments have been used to extract the thermodynamics and kinetics of folding and unfolding reactions. In hopping experiments, a molecule executes transitions between the unfolded and folded states at a preset value of the force [constant force mode (CFM) under force feedback] or trap position [passive mode (PM) without feedback] and the force-dependent kinetic rates extracted from the lifetime of each state (CFM) and the rupture force distributions (PM) using the Bell-Evans model. However, hopping experiments in the CFM are known to overestimate molecular distances and folding free energies for fast transitions compared to the response time of the feedback. In contrast, kinetic rate measurements from pulling experiments have been mostly done in the PM while the CFM is seldom implemented in pulling protocols. Here, we carry out hopping and pulling experiments in a short DNA hairpin in the PM and CFM at three different temperatures (6 °C, 25 °C, and 45 °C) exhibiting largely varying kinetic rates. As expected, we find that equilibrium hopping experiments in the CFM and PM perform well at 6 °C (where kinetics are slow), whereas the CFM overestimates molecular parameters at 45 °C (where kinetics are fast). In contrast, nonequilibrium pulling experiments perform well in both modes at all temperatures. This demonstrates that the same kind of feedback algorithm in the CFM leads to more reliable determination of the folding reaction parameters in irreversible pulling experiments.
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28 March 2018
Research Article|
February 01 2018
Force feedback effects on single molecule hopping and pulling experiments
Special Collection:
Single Molecule Biophysics
M. Rico-Pasto;
M. Rico-Pasto
1
Departament de Fisica de la Materia Condensada, Universitat de Barcelona
, C/ Marti i Franques 1, 08028 Barcelona, Spain
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I. Pastor;
I. Pastor
1
Departament de Fisica de la Materia Condensada, Universitat de Barcelona
, C/ Marti i Franques 1, 08028 Barcelona, Spain
2
CIBER_BNN, Instituto de Salud Carlos III
, 28029 Madrid, Spain
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F. Ritort
F. Ritort
1
Departament de Fisica de la Materia Condensada, Universitat de Barcelona
, C/ Marti i Franques 1, 08028 Barcelona, Spain
2
CIBER_BNN, Instituto de Salud Carlos III
, 28029 Madrid, Spain
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J. Chem. Phys. 148, 123327 (2018)
Article history
Received:
October 24 2017
Accepted:
January 15 2018
Citation
M. Rico-Pasto, I. Pastor, F. Ritort; Force feedback effects on single molecule hopping and pulling experiments. J. Chem. Phys. 28 March 2018; 148 (12): 123327. https://doi.org/10.1063/1.5010303
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