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|Title:||Mechanical characterization of protein L in the low-force regime by electromagnetic tweezers/evanescent nanometry|
|Authors:||Liu, R. |
|Source:||Liu, R., Garcia-Manyes, S., Sarkar, A., Badilla, C.L., Fernández, J.M. (2009). Mechanical characterization of protein L in the low-force regime by electromagnetic tweezers/evanescent nanometry. Biophysical Journal 96 (9) : 3810-3821. ScholarBank@NUS Repository. https://doi.org/10.1016/j.bpj.2009.01.043|
|Abstract:||Mechanical manipulation at the single molecule level of proteins exhibiting mechanical stability poses a technical challenge that has been almost exclusively approached by atomic force microscopy (AFM) techniques. However, due to mechanical drift limitations, AFM techniques are restricted to experimental recordings that last less than a minute in the highforce regime. Here we demonstrate a novel combination of electromagnetic tweezers and evanescent nanometry that readily captures the forced unfolding trajectories of protein L at pulling forces as low as 10 ∼ 15 pN. Using this approach, we m onitor unfolding and refolding cycles of the same polyprotein for a period of time longer than 30 min. From such long-lasting recordings, we obtain ensemble averages of unfolding step sizes and rates that are consistent with single-molecule AFM data obtained at higher stretching forces. The unfolding kinetics of protein L at low stretching forces confirms and extends the observations that the mechanical unfolding rate is exponentially dependent on the pulling force within a wide range of stretching forces spanning from 13 pN up to 120 pN. Our experiments demonstrate a novel approach for the mechanical manipulation of single proteins for extended periods of time in the low-force regime. © 2009 by the Biophysical Society.|
|Source Title:||Biophysical Journal|
|Appears in Collections:||Staff Publications|
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