Tracking unfolding and refolding reactions of single proteins using atomic force microscopy methods

Bujalowski, Paul J.; Oberhauser, Andrés F.

doi:10.1016/j.ymeth.2013.03.010

Cited by 30 publications

(24 citation statements)

References 115 publications

(145 reference statements)

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“…During the recent decades, it has been shown convincingly that osmoprotectors function by promoting protein refolding and suppressing aggregation of different proteins, both in vitro and in vivo [1,9,20,21,[62][63][64][65][66][67][68][69][70][71][72][73][74][75][76][77]. It should be noted that osmolytes may be capable of regulating the activity of the molecular chaperones GroEL, DnaK, and ClpB in a concentration-dependent manner [78][79][80].…”

Section: Protein Folding In Osmolyte Solutionsmentioning

confidence: 99%

Protein folding and stability in the presence of osmolytes

et al. 2016

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Osmolytes are molecules whose function, among others, is to balance the hydrostatic pressure between the intracellular and extracellular compartments. Accumulation of osmolytes in a cell occurs in response to stress caused by changes in pressure, temperature, pH, or the concentration of inorganic salts. Osmolytes can prevent the denaturation of native proteins and promote the renaturation of unfolded proteins. Investigation of the roles of osmolyte in these processes is essential for our understanding of the mechanisms of protein folding and function in vivo. The large number of published reports that have been devoted to the effects of osmolytes on proteins are not always consistent with each other. In this review, an attempt is made to systemize the array of data on this subject and to consider the problem of protein folding and stability in osmolyte solutions from a single viewpoint.

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Section: Protein Folding In Osmolyte Solutionsmentioning

confidence: 99%

Protein folding and stability in the presence of osmolytes

et al. 2016

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“…Over approximately the past decade, static force‐extension measurements of single polymer chains have been extensively performed by many researchers . Many articles have already been published on the use of SMFS for characterizing polymers and bio‐macromolecules, such as protein folding, elasticity of macromolecules, DNA mechanics, and the identification of individual molecules . Herein, our focus is on analyzing the physical properties of single polymer chains.…”

Section: Introductionmentioning

confidence: 99%

Investigating the Dynamic Viscoelasticity of Single Polymer Chains using Atomic Force Microscopy

Liang

Nakajima

2019

J Polym Sci B Polym Phys

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In single‐molecule force spectroscopy (SMFS), many studies have focused on the elasticity and conformation of polymer chains, but little attention has been devoted to the dynamic properties of single polymer chains. In this study, we measured the energy dissipation and elastic properties of single polystyrene (PS) chains in toluene, methanol, and N,N‐dimethylformamide using a homemade piezo‐control and data acquisition system externally coupled to a commercial atomic force microscope (AFM), which provided more accurate information regarding the dynamic properties of the PS chains. We quantitatively measured the chain length‐dependent changes in the stiffness and viscosity of a single chain using a phenomenological model consistent with the theory of viscoelasticity for polymer chains in dilute solution. The effective viscosity of a polymer chain can be determined using the Kirkwood model, which is independent of the intrinsic viscosity of the solvent and dependent on the interaction between the polymer and solvent. The results indicated that the viscosity of a single PS chain is dominated by the interaction between the polymer and solvent. © 2019 Wiley Periodicals, Inc. J. Polym. Sci., Part B: Polym. Phys. 2019, 57, 1736–1743

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“…A wide range of biophysical techniques such as small-angle x-ray scattering (SAXS), 6,7 NMR spectroscopy, 8,9 hydrogen deuterium exchange (HDX), 10 mass spectrometry 11,12 and circular dichroism 13 have been employed to study protein folding and unfolding pathways. A range of methods such as thermally induced, [6][7][8][9]12 radiation induced, 14,15 acid induced 8,10,16,17 and mechanical 18 unfolding have also been employed to follow unfolding pathways. In rapid development since the 1970's, the hybrid technique ion mobility-mass spectrometry (IM-MS) has evolved as a powerful and sensitive gas-phase technique able to elucidate information regarding protein conformational changes.…”

Section: Introductionmentioning

confidence: 99%

Insights into the Conformations of Three Structurally Diverse Proteins: Cytochrome c, p53, and MDM2, Provided by Variable-Temperature Ion Mobility Mass Spectrometry

et al. 2015

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Thermally induced conformational transitions of three proteins of increasing intrinsic disorder-cytochrome c, the tumor suppressor protein p53 DNA binding domain (p53 DBD), and the N-terminus of the oncoprotein murine double minute 2 (NT-MDM2)-have been studied by native mass spectrometry and variable-temperature drift time ion mobility mass spectrometry (VT-DT-IM-MS). Ion mobility measurements were carried out at temperatures ranging from 200 to 571 K. Multiple conformations are observable over several charge states for all three monomeric proteins, and for cytochrome c, dimers of significant intensity are also observed. Cytochrome c [M + 5H](5+) ions present in one conformer of CCS ∼1200 Å(2), undergoing compaction in line with the reported Tmelt = 360.15 K before slight unfolding at 571 K. The more extended [M + 7H](7+) cytochrome c monomer presents as two conformers undergoing similar compaction and structural rearrangements, prior to thermally induced unfolding. The [D + 11H](11+) dimer presents as two conformers, which undergo slight structural compaction or annealing before dissociation. p53 DBD follows a trend of structural collapse before an increase in the observed collision cross section (CCS), akin to that observed for cytochrome c but proceeding more smoothly. At 300 K, the monomeric charge states present in two conformational families, which compact to one conformer of CCS ∼1750 Å(2) at 365 K, in line with the low solution Tmelt = 315-317 K. The protein then extends to produce either a broad unresolved CCS distribution or, for z > 9, two conformers. NT-MDM2 exhibits a greater number of structural rearrangements, displaying charge-state-dependent unfolding pathways. DT-IM-MS experiments at 200 K resolve multiple conformers. Low charge state species of NT-MDM2 present as a single compact conformational family centered on CCS ∼1250 Å(2) at 300 K. This undergoes conformational tightening in line with the solution Tmelt = 348 K before unfolding at the highest temperatures. The more extended charge states present in two or more conformers at room temperature, undergoing thermally induced unfolding before significant structural collapse or annealing at high temperatures. Variable-temperature IM-MS is here shown to be an exciting approach to discern protein unfolding pathways for conformationally diverse proteins.

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Tracking unfolding and refolding reactions of single proteins using atomic force microscopy methods

Cited by 30 publications

References 115 publications

Protein folding and stability in the presence of osmolytes

Protein folding and stability in the presence of osmolytes

Investigating the Dynamic Viscoelasticity of Single Polymer Chains using Atomic Force Microscopy

Insights into the Conformations of Three Structurally Diverse Proteins: Cytochrome c, p53, and MDM2, Provided by Variable-Temperature Ion Mobility Mass Spectrometry

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