Calibration of a dual-trap optical tweezers for single molecule force spectroscopy study

2015 ◽  
Author(s):  
Guoqing Wang ◽  
Chunguang Hu ◽  
Xiaoqing Gao ◽  
Chenguang Su ◽  
Sirong Wang ◽  
...  
Keyword(s):  
Optical Tweezers ◽  
Single Molecule ◽  
Force Spectroscopy ◽  
Spectroscopy Study ◽  
Single Molecule Force Spectroscopy

scholarly journals Single-molecule force spectroscopy reveals folding steps associated with hormone binding and activation of the glucocorticoid receptor

2018 ◽  
Vol 115 (46) ◽  
pp. 11688-11693 ◽  
Author(s):  
Thomas Suren ◽  
Daniel Rutz ◽  
Patrick Mößmer ◽  
Ulrich Merkel ◽  
Johannes Buchner ◽  
...  
Keyword(s):  
Free Energy ◽  
Glucocorticoid Receptor ◽  
Ligand Binding ◽  
Optical Tweezers ◽  
Single Molecule ◽  
Mechanical Load ◽  
Force Spectroscopy ◽  
Folding Pathway ◽  
Hormone Binding ◽  
Single Molecule Force Spectroscopy

The glucocorticoid receptor (GR) is a prominent nuclear receptor linked to a variety of diseases and an important drug target. Binding of hormone to its ligand binding domain (GR-LBD) is the key activation step to induce signaling. This process is tightly regulated by the molecular chaperones Hsp70 and Hsp90 in vivo. Despite its importance, little is known about GR-LBD folding, the ligand binding pathway, or the requirement for chaperone regulation. In this study, we have used single-molecule force spectroscopy by optical tweezers to unravel the dynamics of the complete pathway of folding and hormone binding of GR-LBD. We identified a “lid” structure whose opening and closing is tightly coupled to hormone binding. This lid is located at the N terminus without direct contacts to the hormone. Under mechanical load, apo-GR-LBD folds stably and readily without the need of chaperones with a folding free energy of 41 kBT (24 kcal/mol). The folding pathway is largely independent of the presence of hormone. Hormone binds only in the last step and lid closure adds an additional 12 kBT of free energy, drastically increasing the affinity. However, mechanical double-jump experiments reveal that, at zero force, GR-LBD folding is severely hampered by misfolding, slowing it to less than 1·s−1. From the force dependence of the folding rates, we conclude that the misfolding occurs late in the folding pathway. These features are important cornerstones for understanding GR activation and its tight regulation by chaperones.

scholarly journals Single molecule force spectroscopy studies of DNA denaturation by T4 gene 32 protein

2004 ◽  
Vol 18 (2) ◽  
pp. 203-211 ◽  
Author(s):  
Mark C. Williams ◽  
Kiran Pant ◽  
Ioulia Rouzina ◽  
Richard L. Karpel
Keyword(s):  
Dna Binding ◽  
Optical Tweezers ◽  
Single Molecule ◽  
Protein Interactions ◽  
Force Spectroscopy ◽  
Melting Transition ◽  
Dna Molecules ◽  
Force Extension ◽  
Single Molecule Force Spectroscopy ◽  
Gene 32

Single molecule force spectroscopy is an emerging technique that can be used to measure the biophysical properties of single macromolecules such as nucleic acids and proteins. In particular, single DNA molecule stretching experiments are used to measure the elastic properties of these molecules and to induce structural transitions. We have demonstrated that double‒stranded DNA molecules undergo a force‒induced melting transition at high forces. Force–extension measurements of single DNA molecules using optical tweezers allow us to measure the stability of DNA under a variety of solution conditions and in the presence of DNA binding proteins. Here we review the evidence of DNA melting in these experiments and discuss the example of DNA force‒induced melting in the presence of the single‒stranded DNA binding protein T4 gene 32. We show that this force spectroscopy technique is a useful probe of DNA–protein interactions, which allows us to obtain binding rates and binding free energies for these interactions.

scholarly journals Single-Molecule Force Spectroscopy Study on Modular Resilin Fusion Protein

ACS Omega ◽  
2017 ◽  
Vol 2 (10) ◽  
pp. 6906-6915 ◽  
Author(s):  
Alessandra Griffo ◽  
Hendrik Hähl ◽  
Samuel Grandthyll ◽  
Frank Müller ◽  
Arja Paananen ◽  
...  
Keyword(s):  
Fusion Protein ◽  
Single Molecule ◽  
Force Spectroscopy ◽  
Spectroscopy Study ◽  
Single Molecule Force Spectroscopy

scholarly journals Single-Molecule Force Spectroscopy Study on the Mechanism of RNA Disassembly in Tobacco Mosaic Virus

2013 ◽  
Vol 105 (12) ◽  
pp. 2790-2800 ◽  
Author(s):  
Ningning Liu ◽  
Ying Chen ◽  
Bo Peng ◽  
Yuan Lin ◽  
Qian Wang ◽  
...  
Keyword(s):  
Tobacco Mosaic Virus ◽  
Mosaic Virus ◽  
Single Molecule ◽  
Force Spectroscopy ◽  
Spectroscopy Study ◽  
Single Molecule Force Spectroscopy

Single-molecule-force spectroscopy study of the mechanism of interactions between TSP-1 and CD47

2014 ◽  
Vol 57 (12) ◽  
pp. 1716-1722 ◽  
Author(s):  
YanGang Pan ◽  
Feng Wang ◽  
YanHou Liu ◽  
Yong-Guang Yang ◽  
HongDa Wang
Keyword(s):  
Single Molecule ◽  
Force Spectroscopy ◽  
Spectroscopy Study ◽  
Single Molecule Force Spectroscopy
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