scholarly journals Gas Phase Stability of Protein Ions in a Cyclic Ion Mobility Spectrometry Travelling Wave Device

2018 ◽  
Author(s):  
Charles Eldrid ◽  
Jakub Ujma ◽  
Symeon Kalfas ◽  
nick tomczyk ◽  
Kevin Giles ◽  
...  
Keyword(s):  
Cytochrome C ◽  
Gas Phase ◽  
Ion Mobility ◽  
Structural Changes ◽  
Structural Information ◽  
Travelling Wave ◽  
Resolving Power ◽  
Small Proteins ◽  
Multimeric Complex ◽  
Protein Ions

<div>Ion mobility mass spectrometry (IM-MS) allows separation of native protein ions into “conformational families”. Increasing the IM resolving power should allow finer structural information to be obtained, and can be achieved by increasing the length of the IM separator. This, however, increases the time that protein ions spend in the gas phase and previous experiments have shown that the initial conformations of small proteins can be lost within tens of milliseconds. Here, we report on investigations of protein ion stability using a multi-pass travelling wave (TW) cyclic IM (cIM) device. Using this device, minimal structural changes were observed for Cytochrome C after hundreds of milliseconds, while no changes were observed for a larger multimeric complex (Concanavalin A). The geometry of the instrument (Q-cIM-ToF) also enables complex tandem IM experiments to be performed which were used to obtain more detailed collision induced unfolding pathways for Cytochrome C. The novel instrument geometry provide unique capabilities with the potential to expand the field of protein analysis via IM-MS.</div>

scholarly journals Gas Phase Stability of Protein Ions in a Cyclic Ion Mobility Spectrometry Travelling Wave Device

2018 ◽  
Author(s):  
Charles Eldrid ◽  
Jakub Ujma ◽  
Symeon Kalfas ◽  
nick tomczyk ◽  
Kevin Giles ◽  
...  
Keyword(s):  
Cytochrome C ◽  
Gas Phase ◽  
Ion Mobility ◽  
Structural Changes ◽  
Structural Information ◽  
Travelling Wave ◽  
Resolving Power ◽  
Small Proteins ◽  
Multimeric Complex ◽  
Protein Ions

<div>Ion mobility mass spectrometry (IM-MS) allows separation of native protein ions into “conformational families”. Increasing the IM resolving power should allow finer structural information to be obtained, and can be achieved by increasing the length of the IM separator. This, however, increases the time that protein ions spend in the gas phase and previous experiments have shown that the initial conformations of small proteins can be lost within tens of milliseconds. Here, we report on investigations of protein ion stability using a multi-pass travelling wave (TW) cyclic IM (cIM) device. Using this device, minimal structural changes were observed for Cytochrome C after hundreds of milliseconds, while no changes were observed for a larger multimeric complex (Concanavalin A). The geometry of the instrument (Q-cIM-ToF) also enables complex tandem IM experiments to be performed which were used to obtain more detailed collision induced unfolding pathways for Cytochrome C. The novel instrument geometry provide unique capabilities with the potential to expand the field of protein analysis via IM-MS.</div>

scholarly journals Multiplexed Conformationally-Selective, Localized Gas-Phase Hydrogen Deuterium Exchange of Protein Ions Enabled by Transmission-Mode Electron Capture Dissociation

2021 ◽  
Author(s):  
Ritu Chaturvedi ◽  
Ian Webb
Keyword(s):  
Cytochrome C ◽  
Gas Phase ◽  
Charge State ◽  
Electron Capture ◽  
Electron Capture Dissociation ◽  
Ion Mobility ◽  
Transmission Mode ◽  
Hydrogen Deuterium Exchange ◽  
Protein Ions ◽  
Hydrogen Deuterium

In this article, we present an approach for conformationally multiplexed localized hydrogen deuterium exchange (HDX) of gas-phase protein ions facilitated by ion mobility (IM) followed by electron capture dissociation (ECD). A quadrupole-ion mobility-time of flight instrument previously modified to enable ECD in transmission mode (without ion trapping) immediately following a mobility separation was further modified to allow for deuterated ammonia (ND3) to be leaked in after m/z selection. Collisional activation was minimized to prevent deuterium scrambling from giving structurally irrelevant results. This arrangement was demonstrated with the extensively studied protein folding models ubiquitin and cytochrome c. Ubiquitin was ionized from conditions that stabilize the native state and conditions that stabilize the partially-folded A-state. IM of deuterated ubiquitin 6+ ions allowed the separation of more compact conformers from more extended conformers. ECD of the separated subpopulations revealed that the more extended (later arriving) conformers had significant, localized differences in the amount of HDX observed. The 5+ charge state showed greater protection against HDX than the compact 6+ conformer, and the 11+ charge state, ionized from conditions that stabilize the A-state, showed much greater deuterium incorporation. The 7+ ions of cytochrome c ionized from aqueous conditions showed greater HDX with exterior and more unstructured regions of the protein, while interior, structured regions, especially those involved in heme binding, were more protected against exchange. These results, as well as potential future methods and experiments, are discussed herein.

scholarly journals Gas Phase Stability of Protein Ions in a Cyclic Ion Mobility Spectrometry Traveling Wave Device

2019 ◽  
Vol 91 (12) ◽  
pp. 7554-7561 ◽  
Author(s):  
Charles Eldrid ◽  
Jakub Ujma ◽  
Symeon Kalfas ◽  
Nick Tomczyk ◽  
Kevin Giles ◽  
...  
Keyword(s):  
Gas Phase ◽  
Ion Mobility Spectrometry ◽  
Phase Stability ◽  
Ion Mobility ◽  
Traveling Wave ◽  
Protein Ions ◽  
Wave Device

scholarly journals Towards a hand-held, fast, and sensitive gas chromatograph-ion mobility spectrometer for detecting volatile compounds

2020 ◽  
Author(s):  
André Ahrens ◽  
Stefan Zimmermann
Keyword(s):  
Gas Phase ◽  
Ion Mobility ◽  
Atmospheric Pressure ◽  
Response Times ◽  
Atmospheric Pressure Chemical Ionization ◽  
Real Life ◽  
High Sensitivity ◽  
Resolving Power ◽  
Gas Chromatograph ◽  
Ionization Processes

AbstractIon mobility spectrometers can detect gaseous compounds at atmospheric pressure in the range of parts per trillion within a second. Due to their fast response times, high sensitivity, and limited instrumental effort, they are used in a variety of applications, especially as mobile or hand-held devices. However, most real-life samples are gas mixtures, which can pose a challenge for IMS with atmospheric pressure chemical ionization mainly due to competing gas-phase ionization processes. Therefore, we present a miniaturized drift tube IMS coupled to a compact gas chromatograph for pre-separation, built of seven bundled standard GC columns (Rtx-Volatiles, Restek GmbH) with 250 μm ID and 1.07 m in length. Such pre-separation significantly reduces chemical cross sensitivities caused by competing gas-phase ionization processes and adds orthogonality. Our miniaturized GC-IMS system is characterized with alcohols, halocarbons, and ketones as model substances, reaching detection limits down to 70 pptv with IMS averaging times of just 125 ms. It separates test mixtures of ketones and halocarbons within 180 s and 50 s, respectively. The IMS has a short drift length of 40.6 mm and reaches a high resolving power of RP = 68.

scholarly journals Effective Discrimination of Gas-Phase Peptide Conformers using TIMS-ECD-ToF MS/MS

2021 ◽  
Author(s):  
Kevin Jeanne dit fouque ◽  
Malte Wellmann ◽  
Dennys Leyva ◽  
Miguel Santos-Fernandez ◽  
Yarixa Cintron Diaz ◽  
...  
Keyword(s):  
Substance P ◽  
Gas Phase ◽  
Ion Mobility ◽  
Structural Information ◽  
Angiotensin I ◽  
Tof Ms ◽  
Model Peptides

In the present work, four, well-studied, model peptides (e.g., substance P, bradykinin, angiotensin I and AT-Hook 3) were used to correlate structural information provided by ion mobility and ECD/CID fragmentation...

scholarly journals Conformational footprinting of proteins using a combination of top-down electron transfer dissociation and ion mobility

2018 ◽  
Author(s):  
Albert Konijnenberg ◽  
Jinyu Li ◽  
Johny Habchi ◽  
Marion Dosnon ◽  
Giulia Rossetti ◽  
...  
Keyword(s):  
Protein Structure ◽  
Gas Phase ◽  
Ion Mobility ◽  
Measles Virus ◽  
Electron Transfer Dissociation ◽  
Structural Information ◽  
Native Mass Spectrometry ◽  
Top Down ◽  
Wide Range ◽  
Protein Ensembles

AbstractIn recent years native mass spectrometry has been increasingly employed to study protein structure. As such a thorough understanding of the effect of the gas-phase on protein structure is becoming increasingly important. We show how a combination of top-down ETD and ion mobility can be used to probe the gas-phase structure of heterogeneous protein ensembles. By applying collisional activation to the non-covalently bound ETD products after IM separation, the peptide fragments can be released while maintaining the conformational information of the protein ion. We studied the unknown gas-phase structures of the measles virus (MeV) phosphoprotein X domain (PXD), which shows a wide range of different conformations in the gas-phase. We then generated structural models by state-of-the-art gas-phase steered molecular dynamics, which we verified using restraints from ion mobility and the fragment patterns observed. Our findings illustrate the applicability of ETD for obtaining conformational specific structural information on heterogeneous protein ensembles.

scholarly journals Protein Unfolding in Freeze Frames: Intermediates of Ubiquitin and Lysozyme Revealed by Variable Temperature Ion Mobility-Mass Spectrometry

2020 ◽  
Author(s):  
Jakub Ujma ◽  
Jacquelyn Jhingree ◽  
Rosie Upton ◽  
Florian Benoit ◽  
Bruno Bellina ◽  
...  
Keyword(s):  
Ion Mobility ◽  
Protein Unfolding ◽  
Collision Cross Section ◽  
Activation Barrier ◽  
Variable Temperature ◽  
Activation Process ◽  
Small Proteins ◽  
Protein Ions ◽  
New Instrument ◽  
Native Fold

As experimentalists, we normally rely on assessing observables. However sometimes, the most fascinating phenomena are not noticeable directly. An example of such is our data and the corresponding interpretation presented in this manuscript. We have designed and constructed a ion mobility mass spectrometer (acs.analchem.6b01812) capable of taking ion mobility measurements over a temperature range from 150-500K. We chose to benchmark this new instrument, using the small proteins Ubiquitin and Lysozyme extensively studied as a “model proteins” in many in-silico, -solution and -vacuo studies focusing on conformational dynamics.In this work, we activate and subsequently thermally equilibrate the protein ions at several temperatures prior to collision cross section measurement. For Ubiquitin at 300K and above, the protein unfolds in a “step-wise” fashion as previously reported (by David Clemmer) and for other proteins including lysozyme, and cytochrome c by us and also by Martin Jarrold. However, to our surprise, activation and equilibration of ubiquitin at 150K yields a plethora of highly extended forms of the protein. We attribute these as kinetically trapped unfolding intermediates. Since the activation process is the same at both temperatures we infer that the unfolding must always proceed via these extended intermediate forms, which then converge to commonly reported conformations. Intriguingly, this “convergence” appears to occur mostly below the temperature of irreversible conformational thermal transition of Ubiquitin reported in many solution phase studies. For Lysozyme the same experiment is performed and similar results are obtained although we cannot activate the gaseous ensemble too far from the native fold and the activation barrier to refolding is low enough to allow it to be re-accessed on the experimental timescale we use.

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