Chemical, Thermal, and Electric Field Induced Unfolding of Single Protein Molecules Studied Using Nanopores

Freedman, Kevin J.; Jürgens, Maike C.; Prabhu, Anmiv S.; Ahn, Chi Won; Jemth, Per; Edel, Joshua B.; Kim, Min Jun

doi:10.1021/ac2001725

Cited by 127 publications

(144 citation statements)

References 40 publications

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“…It is puzzling why the native state BSA in a nanopore produced such a broad peak in ΔI b , implying a broad distribution of the excluded volume Λ and translocation configurations. Broad range of ΔI b distributions were also observed by previous reports studying BSA and other protein translocation in solid-state nanopores [9, 16–19, 21, 25–27], suggesting some protein molecules going through solid-state nanopores with multiple conformations, some could be partially denatured.…”

Section: Discussionsupporting

confidence: 76%

“…However, this estimation has ignored complicated issues such as the space inside a folded protein, hydration layer and territory bound water and ions on protein surface, the real excluded volume of a protein molecule in a nanopore is expected to be larger. For example, here we calculate Λ fold =87.0 nm 3 for a folded BSA, this is smaller than the value reported by Freedman et al [25] calculated by formula Λ=(4/3)πa 2 b =117 nm 3 using the dimensions shown in Fig. 1d, and it is also smaller than experimentally measured value166±29 nm 3 .…”

Section: Principles Of Measuring Protein Unfolding By a Solid-statcontrasting

confidence: 79%

“…Solid-state nanopores fabricated with desired geometry and dimensions, have been demonstrated to be capable of detecting protein molecules both in their native (folded) [16–25] and in their denatured (unfolded) states [9, 26, 27]. Motivated to develop a solid-state nanopore based device to identify proteins by measuring them both in their folded and unfolded states, we have studied the process of protein unfolding with sold-state nanopores.…”

Section: Introductionmentioning

confidence: 99%

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Characterization of Protein Unfolding with Solid-state Nanopores

Li¹,

Fologea²,

Rollings³

et al. 2014

PPL

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In this work, we review the process of protein unfolding characterized by a solid-state nanopore based device. The occupied or excluded volume of a protein molecule in a nanopore depends on the protein’s conformation or shape. A folded protein has a larger excluded volume in a nanopore thus it blocks more ionic current flow than its unfolded form and produces a greater current blockage amplitude. The time duration a protein stays in a pore also depends on the protein’s folding state. We use Bovine Serum Albumin (BSA) as a model protein to discuss this current blockage amplitude and the time duration associated with the protein unfolding process. BSA molecules were measured in folded, partially unfolded, and completely unfolded conformations in solid-state nanopores. We discuss experimental results, data analysis, and theoretical considerations of BSA protein unfolding measured with silicon nitride nanopores. We show this nanopore method is capable of characterizing a protein’s unfolding process at single molecule level. Problems and future studies in characterization of protein unfolding using a solid-state nanopore device will also be discussed.

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Section: Discussionsupporting

confidence: 76%

Section: Principles Of Measuring Protein Unfolding By a Solid-statcontrasting

confidence: 79%

Section: Introductionmentioning

confidence: 99%

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Characterization of Protein Unfolding with Solid-state Nanopores

Li¹,

Fologea²,

Rollings³

et al. 2014

PPL

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“…In the past decade, synthetic nanopores have been widely used for single molecule detection [2,[15][16][17] [17], and e-field can influence interactions between the protein and pore which can trigger protein denaturation inside a nanopore [33]. Thus, a comprehensive understanding of the conformational changes is important because of the nature of detection using ionic current measurement.…”

Section: Introductionmentioning

confidence: 99%

Molecular Dynamics Study of Protein Deformation through Solid-State Nanopore

Hasan¹,

Mahmood²,

Khanzada³

et al. 2016

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Electrophoretic translocation through solid-state nanopores is a promising technique for identification of specific proteins and protein complexes. In a typical protein detection experiment, an external electric field (e-field) is applied across the nanopore and the ionic current is measured while the molecule passes through the pore. It is very important that the protein retains its structure and functionality under applied e-field and experimental conditions. However, the elongation and deformation of the 3D structure may bias the detection scheme. This work presents a theoretical assessment of protein deformation in a nanopore experiment due to applied e-field. We used nanoscale molecular dynamics (NAMD) simulations to investigate the deformation of a model protein during translocation through a silicon nitride nanopore in an aqueous solution under varying e-fields. We simulated the experimental conditions and performed quantitative analysis of the variations in thrombin protein's 3D structure. No mechanical force was applied on the protein to ensure the sole contribution of e-field on the deformation. The conformational changes of thrombin's structure were quantified in terms of root mean square deviation (RMSD) and radial distribution function. It was observed that large voltages resulted in deformations in protein structure. The protein molecule got stretched under the influence of high e-field as it traveled through nanopore. The interaction mechanism between nanopore and protein is crucial to understand the detection dynamics. The results can guide better design of experiments with proteins confined in a nanopore.

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“…It is well known that intense electric fields can profoundly affect the physical properties and reactivity of dissolved molecules and, based on experimental results, they have been proven to play an essential role in proteins behavior, including folding, molecular recognition, and catalytic functions [14,15]. Recently, MD technique has been suggested as the best approach for shedding light on the different hypotheses on the interaction between electromagnetic fields and biological targets [16,17], for which a complete theoretical investigation is still lacking.In particular fully atomistic simulations have proven to be a valuable methodology: to study the effect of microwaves on enzymes in solution [18,19], to investigate the action of pulsed electric fields on biochemical reactions in confined environments with outcomes confirmed by the experimental data [20] and to provide insight in the dynamic behavior of small peptides dispersed in water solutions, under the influences of external electric fields such as those exerted by biomedical sensors and/or field-effect transistors [21].…”

Section: Introductionmentioning

confidence: 99%

Evaluation of Protein Electrostatic Potential from Molecular Dynamics Simulations in the Presence of Exogenous Electric Fields: The Case Study of Myoglobin

Marracino

Casciola

Liberti

et al. 2014

Computational Electrostatics for Biological Applications

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When studying proteins in solution it is apparent that electrostatic interactions play a role in folding, conformational stability, and other chemicalphysical properties. Electrostatics considers the evaluation of the static electrical field that is formed between charged species once a rearrangement of their charge distributions has occurred due to the influence of each other and their local environment. A powerful tool used to follow the many interactions among the polar and/or charged residues is computer simulations, which can provide atomic-scale information on energetic and dynamic contributions of the bio-molecular structure. Here we use molecular dynamics (MD) simulations to map on a three-dimensional space the electrostatic interactions within the protein itself and of the protein with its aqueous environment. The method has been first tested on a simulation domain of water molecules and then applied to the myoglobin-water system. The presence of intense electric fields has also been considered and some representative results are discussed. IntroductionThe electrostatic interactions between charged atoms in natural proteins play a central role in specifying protein topology, modulating stability of the molecule, and allowing for the important catalytic properties of enzymes. Such kind of interactions are at the basis of molecular characterization; when studying biomolecules it is becoming increasingly evident that electrostatic interactions play a role in folding, conformational stability, enzyme activity, and binding energies as well as in

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Chemical, Thermal, and Electric Field Induced Unfolding of Single Protein Molecules Studied Using Nanopores

Cited by 127 publications

References 40 publications

Characterization of Protein Unfolding with Solid-state Nanopores

Characterization of Protein Unfolding with Solid-state Nanopores

Molecular Dynamics Study of Protein Deformation through Solid-State Nanopore

Evaluation of Protein Electrostatic Potential from Molecular Dynamics Simulations in the Presence of Exogenous Electric Fields: The Case Study of Myoglobin

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