Two energy barriers and a transient intermediate state determine the unfolding and folding dynamics of cold shock protein

AbstractCold shock protein (Csp) is a typical two-state folding model protein which has been widely studied by biochemistry and single molecule techniques. Recently two-state property of Csp was confirmed by atomic force microscopy (AFM) through direct pulling measurement, while several long-lifetime intermediate states were found by force-clamp AFM. We systematically studied force-dependent folding and unfolding dynamics of Csp using magnetic tweezers with intrinsic constant force capability. Here we report that Csp mostly folds and unfolds with a single step over force range from 5 pN to 50 pN, and the unfolding rates show different force sensitivities at forces below and above ~8 pN, which determines a free energy landscape with two barriers and a transient intermediate state between them along one transition pathway. Our results provide a new insight on protein folding mechanism of two-state proteins.

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Free energy landscape with two barriers and a transient intermediate state determining the unfolding and folding dynamics of cold shock protein

10.21203/rs.3.rs-449394/v1 ◽

2021 ◽

Author(s):

Haiyan Hong ◽

Zilong Guo ◽

Hao Sun ◽

Ping Yu ◽

Huanhuan Su ◽

...

Keyword(s):

Free Energy ◽

Single Molecule ◽

Cold Shock ◽

Energy Landscape ◽

Magnetic Tweezers ◽

Single Step ◽

Cold Shock Protein ◽

Free Energy Landscape ◽

Transient Intermediate ◽

Two Barriers

Abstract Cold shock protein (Csp) is a typical two-state folding model protein which has been widely studied by biochemistry and single molecule techniques. Recently two-state property of Csp was confirmed by atomic force microscopy (AFM) through direct pulling measurement, while several long-lifetime intermediate states were found by force-clamp AFM. We systematically studied force-dependent folding and unfolding dynamics of Csp using magnetic tweezers with intrinsic constant force capability. We found that Csp mostly folds and unfolds with a single step over force range from 5 pN to 50 pN, and the unfolding rates show different force sensitivities at forces below and above ~ 8 pN, which determines a free energy landscape with two barriers and a transient intermediate between them along one transition pathway. Our results provide a new insight on protein folding mechanism of two-state proteins.

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Single molecule FRET investigation of pressure-driven unfolding of cold shock protein A

The Journal of Chemical Physics ◽

10.1063/1.5009662 ◽

2018 ◽

Vol 148 (12) ◽

pp. 123336 ◽

Cited By ~ 1

Author(s):

Sven Schneider ◽

Hauke Paulsen ◽

Kim Colin Reiter ◽

Erik Hinze ◽

Cordelia Schiene-Fischer ◽

...

Keyword(s):

Single Molecule ◽

Cold Shock ◽

Protein A ◽

Cold Shock Protein ◽

Single Molecule Fret ◽

Pressure Driven

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Single-Molecule Force Spectroscopy Identifies a Small Cold Shock Protein as Being Mechanically Robust

The Journal of Physical Chemistry B ◽

10.1021/jp310442s ◽

2013 ◽

Vol 117 (6) ◽

pp. 1819-1826 ◽

Cited By ~ 21

Author(s):

Toni Hoffmann ◽

Katarzyna M. Tych ◽

David J. Brockwell ◽

Lorna Dougan

Keyword(s):

Single Molecule ◽

Cold Shock ◽

Force Spectroscopy ◽

Cold Shock Protein ◽

Single Molecule Force Spectroscopy

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Faculty Opinions recommendation of Cold shock protein 1 chaperones mRNAs during translation in Arabidopsis thaliana.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.718017826.793482971 ◽

2013 ◽

Author(s):

Motoaki Seki ◽

Kentaro Nakaminami

Keyword(s):

Arabidopsis Thaliana ◽

Cold Shock ◽

Cold Shock Protein

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Study on the transgenic tobacco expressing truncated wheat cold shock protein gene TA3-13 and analysis of disease resistance

Hereditas (Beijing) ◽

10.3724/sp.j.1005.2011.00520 ◽

2011 ◽

Vol 33 (5) ◽

pp. 520-526 ◽

Cited By ~ 1

Author(s):

Na LI ◽

Xiu-Zhen DU ◽

Xiao-Mei PAN ◽

Jin-Sheng WANG ◽

Cong-Feng SONG

Keyword(s):

Disease Resistance ◽

Transgenic Tobacco ◽

Cold Shock ◽

Protein Gene ◽

Cold Shock Protein

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Electrostatic stabilization of a thermophilic cold shock protein

Journal of Molecular Biology ◽

10.1006/jmbi.2001.5239 ◽

2002 ◽

Vol 316 (1) ◽

pp. 213 ◽

Cited By ~ 1

Author(s):

D. Perl ◽

F.X. Schmid

Keyword(s):

Cold Shock ◽

Cold Shock Protein ◽

Electrostatic Stabilization

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A simple and versatile design concept for fluorophore derivatives with intramolecular photostabilization

Nature Communications ◽

10.1038/ncomms10144 ◽

2016 ◽

Vol 7 (1) ◽

Cited By ~ 62

Author(s):

Jasper H. M. van der Velde ◽

Jens Oelerich ◽

Jingyi Huang ◽

Jochem H. Smit ◽

Atieh Aminian Jazi ◽

...

Keyword(s):

Single Molecule ◽

Unnatural Amino Acids ◽

Photophysical Properties ◽

Super Resolution ◽

Single Step ◽

General Strategy ◽

Self Healing ◽

Chemical Transformations ◽

Scaffolding Strategy ◽

Organic Fluorophore

Abstract Intramolecular photostabilization via triple-state quenching was recently revived as a tool to impart synthetic organic fluorophores with ‘self-healing’ properties. To date, utilization of such fluorophore derivatives is rare due to their elaborate multi-step synthesis. Here we present a general strategy to covalently link a synthetic organic fluorophore simultaneously to a photostabilizer and biomolecular target via unnatural amino acids. The modular approach uses commercially available starting materials and simple chemical transformations. The resulting photostabilizer–dye conjugates are based on rhodamines, carbopyronines and cyanines with excellent photophysical properties, that is, high photostability and minimal signal fluctuations. Their versatile use is demonstrated by single-step labelling of DNA, antibodies and proteins, as well as applications in single-molecule and super-resolution fluorescence microscopy. We are convinced that the presented scaffolding strategy and the improved characteristics of the conjugates in applications will trigger the broader use of intramolecular photostabilization and help to emerge this approach as a new gold standard.

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