Towards the analysis of high molecular weight proteins and protein complexes using TIMS-MS

Benigni, Paolo; Marin, Rebecca; Molano-Arévalo, Juan Camilo; Garabedian, Alyssa; Wolff, Jeremy J.; Ridgeway, Mark E.; Park, Melvin A.; Fernandez-Lima, Francisco

doi:10.1007/s12127-016-0201-8

Cited by 45 publications

(53 citation statements)

References 123 publications

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“…300 K, at a constant gas flow velocity defined by the pressure difference between entrance funnel P 1 = 1.1–4.3 mbar, and the exit funnel P 2 = 0.6–3.0 mbar. 64, 65 …”

Section: Methodsmentioning

confidence: 99%

Structures of the kinetically trapped i-motif DNA intermediates

Garabedian

Butcher

Lippens

et al. 2016

Phys. Chem. Chem. Phys.

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In the present work, the conformational dynamics and folding pathways of i-motif DNA were studied in solution and in the gas-phase as a function of the solution pH conditions using circular dichroism (CD), photoacoustic calorimetry analysis (PAC), trapped ion mobility spectrometry - mass spectrometry (TIMS-MS), and molecular dynamics (MD). Solution studies showed at thermodynamic equilibrium the existence of a two-state folding mechanism, whereas during the pH = 7.0 → 4.5 transition a fast and slow phase (ΔHfast+ ΔHslow = 43 ± 7 kcal mol−1) with a volume change associated with the formation of hemiprotonated cytosine base pairs and concomitant collapse of the i-motif oligonucleotide into a compact conformation were observed. TIMS-MS experiments showed that gas-phase, kinetically trapped i-motif DNA intermediates produced by nanoESI are preserved, with relative abundances depending on the solution pH conditions. In particular, a folded i-motif DNA structure was observed in nanoESI-TIMS-MS for low charge states in both positive and negative ion mode (e.g., z = +/− 3 to +/−5) at low pH conditions. As solution pH increases, the cytosine deprotonation leads to the loss of cytosine-cytosine+ (C•CH+) base pairing in the CCC strands and in those conditions we observe partially unfolded i-motif DNA conformations in nanoESI-TIMS-MS for higher charge states (e.g., z = − 6 to −9). Collisional induced activation prior TIMS-MS showed the existence of multiple local free energy minima, associated with the i-motif DNA unfolding at z = −6 charge state. For the first time, candidate gas-phase structures are proposed based on mobility measurements of the i-motif DNA unfolding pathway. Moreover, the inspection of partially unfolded i-motif DNA structures (z = −7 and z = −8 charge states) showed that the presence of inner cations may or may not induce conformational changes in the gas-phase. For example, incorporation of ammonium adducts does not lead to major conformational changes while sodium adducts may lead to the formation of sodium mediated bonds between two negatively charged sides inducing the stabilization towards more compact structures in new local, free energy minima in the gas-phase.

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“…300 K, at a constant gas flow velocity defined by the pressure difference between entrance funnel P 1 = 1.1–4.3 mbar, and the exit funnel P 2 = 0.6–3.0 mbar. 64, 65 …”

Section: Methodsmentioning

confidence: 99%

Structures of the kinetically trapped i-motif DNA intermediates

Garabedian

Butcher

Lippens

et al. 2016

Phys. Chem. Chem. Phys.

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“…The data recorded on the trapped IMS system at FSU agrees essentially numerically with these data but only after careful “soft-tuning” of the operating conditions by down tuning the DC and RF electric fields as well as the source conditions (needle potential, desolvation gas temperature) (50). The cross sections of charge states 6,7, and 8 are significantly larger—indicative of substantial collision-induced unfolding of ubiquitin ions during the measurement—when high DC and RF fields are used and the ESI desolvation gas is heated in excess of 370K in a trapped IMS device (54). Charge states 4 and 5, which are not generally observed under native conditions (50,53), should not be considered indicative of the ubiquitin solution structure as cross sections of their “inside-out” gas-phase structures are not distinguishable from that of the ubiquitin solution structure (55).…”

Section: Softness Of Ims-ms Measurements: the Ability To Study Biologmentioning

confidence: 99%

“…(Note that cross sections of “inside-out” structures for charge states +4 or +5 are similar to the solution structure; see reference (55) for details.) Ubiquitin cross sections were taken from references (53), (50), and (54). (B) Trapped IMS spectra for m/z 1060 of bradykinin with and without mass selection in the quadrupole (red and black traces, respectively) comprises monomers and oligomers.…”

Section: Figurementioning

confidence: 99%

The Solution Assembly of Biological Molecules Using Ion Mobility Methods: From Amino Acids to Amyloid β-Protein

Bleiholder

Bowers

2017

Annual Rev. Anal. Chem.

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Ion mobility spectrometry-mass spectrometry (IMS-MS) methods are increasingly used to study noncovalent assemblies of peptides and proteins. This review focuses on the noncovalent self-assembly of amino acids and peptides, systems at the heart of the amyloid process that play a central role in a number of devastating diseases. Three different systems are discussed in detail: the 42-residue peptide amyloid-β42 implicated in the etiology of Alzheimer's disease, several amyloid-forming peptides with 6–11 residues, and the assembly of individual amino acids. We also discuss from a more fundamental perspective the processes that determine how quickly proteins and their assemblies denature when the analyte ion has been stripped of its solvent in an IMS-MS measurement and how to soften the measurement so that biologically meaningful data can be recorded.

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“…36,37 The TIMS devices provide R up to ~400 in a compact form 38 and are readily integrated with various MS platforms, including time-of-flight (ToF) and Fourier Transform MS. 39–42 The TIMS-MS systems have proven useful for rapid separation and structural elucidation of biomolecules, 42–52 for example screening 43 and targeted 40 analysis of complex mixtures, tracking the isomerization kinetics, 44–46 and characterizing the conformational spaces of peptides, 53 DNA, 47 proteins, 54 and macromolecular complexes in native and denatured states. 55 …”

mentioning

confidence: 99%

Fast and Effective Ion Mobility–Mass Spectrometry Separation of d-Amino-Acid-Containing Peptides

et al. 2017

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Despite often minute concentrations in vivo, D-amino acid containing peptides (DAACPs) are crucial to many life processes. Standard proteomics protocols fail to detect them as D/L substitutions do not affect the peptide parent and fragment masses. The differences in fragment yields are often limited, obstructing the investigations of important but low abundance epimers in isomeric mixtures. Separation of D/L-peptides using ion mobility spectrometry (IMS) was impeded by small collision cross section differences (commonly ~1%). Here, broad baseline separation of DAACPs with up to ~30 residues employing trapped IMS with resolving power up to ~340, followed by time-of-flight mass spectrometry is demonstrated. The D/L-pairs co-eluting in one charge state were resolved in another, and epimers merged as protonated species were resolved upon metalation, effectively turning the charge state and cationization mode into extra separation dimensions. Linear quantification down to 0.25% proved the utility of high resolution IMS-MS for real samples with large inter-isomeric dynamic range. Very close relative mobilities found for DAACP pairs using traveling-wave IMS (TWIMS) with different ion sources and faster IMS separations showed the transferability of results across IMS platforms. Fragmentation of epimers can enhance their identification and further improve detection and quantification limits, and we demonstrate the advantages of online mobility separated collision-induced dissociation (CID) followed by high resolution mass spectrometry (TIMS-CID-MS) for epimer analysis.

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Towards the analysis of high molecular weight proteins and protein complexes using TIMS-MS

Cited by 45 publications

References 123 publications

Structures of the kinetically trapped i-motif DNA intermediates

Structures of the kinetically trapped i-motif DNA intermediates

The Solution Assembly of Biological Molecules Using Ion Mobility Methods: From Amino Acids to Amyloid β-Protein

Fast and Effective Ion Mobility–Mass Spectrometry Separation of d-Amino-Acid-Containing Peptides

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