scholarly journals Protein unfolding in freeze frames: intermediate states are revealed by variable temperature ion mobility-mass spectrometry.

2021 ◽  
Author(s):  
Jakub Ujma ◽  
Jacquelyn Jhingree ◽  
Emma Norgate ◽  
Rosie Upton ◽  
Xudong Wang ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Ion Mobility ◽  
Elevated Temperatures ◽  
Alpha Synuclein ◽  
Conformational Space ◽  
Disordered Proteins ◽  
Reduced Form ◽  
Ion Mobility Mass Spectrometry ◽  
Model Framework ◽  
Intermediate States

The gas phase is an idealized laboratory for the study of protein structure, from which it is possible to examine stable and transient forms of mass selected ions in the absence of bulk solvent. With ion mobility-mass spectrometry (IM-MS) apparatus built to operate at both cryogenic and elevated temperatures, we have examined the conformational transitions of the monomeric proteins: ubiquitin, lysozyme and alpha-synuclein as a function of temperature and in source activation. We rationalize the experimental observations with a temperature dependent framework model and comparison to known conformers. Data from ubiquitin shows unfolding transitions that proceed through diverse and highly elongated intermediate states, which converge to more compact structures. These findings contrast with data obtained from lysozyme – a protein where (un)-folding plasticity is restricted by four disulfide linkages, although this is alleviated in its reduced form. For structured proteins, collision activation of the protein ions in- source, enables subsequent “freezing” or thermal annealing of unfolding intermediates whereas disordered proteins restructure substantially at 250 K even without activation, indicating that cold denaturation can occur without solvent. These data are presented in the context of a toy model framework which describes the relative occupancy of the available conformational space.

Ion Mobility Mass Spectrometry Studies of the Inhibition of Alpha Synuclein Amyloid Fibril Formation by ( - )-Epigallocatechin-3-Gallate

2011 ◽  
Vol 64 (1) ◽  
pp. 36 ◽  
Author(s):  
Yanqin Liu ◽  
Lam H. Ho ◽  
John. A. Carver ◽  
Tara L. Pukala
Keyword(s):  
Mass Spectrometry ◽  
Conformational Change ◽  
Ion Mobility ◽  
Amyloid Fibril ◽  
Heterogeneous Systems ◽  
Alpha Synuclein ◽  
Fibril Formation ◽  
Ion Mobility Mass Spectrometry ◽  
Compact Structure ◽  
Amyloid Fibril Formation

Ion mobility-mass spectrometry (IM-MS) is emerging as an important biophysical technique for the structural analysis of proteins and their assemblies, in particular for structurally heterogeneous systems such as those on the protein misfolding and aggregation pathway. Using IM-MS we have monitored amyloid fibril formation of A53T α-synuclein, a mutant synuclein protein associated with Parkinson’s disease, and identified that a conformational change towards a more compact structure occurs during the initial stages of aggregation. Binding of A53T α-synuclein to a flavenoid based amyloid fibril inhibitor, (–)-epigallocatechin-3-gallate, has been observed with a 1:1 stoichiometry. By analysis of ion collision cross-sections, we show epigallocatechin gallate binding prevents protein conformational change, and in turn decreases the formation of fibrillar aggregates.

scholarly journals Ion Mobility Mass Spectrometry Uncovers the Impact of the Patterning of Oppositely Charged Residues on the Conformational Distributions of Intrinsically Disordered Proteins

2018 ◽  
Author(s):  
Rebecca Beveridge ◽  
Lukasz Migas ◽  
Rahul Das ◽  
Rohit Pappu ◽  
Richard Kriwacki ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Ion Mobility ◽  
Intrinsically Disordered Proteins ◽  
Disordered Proteins ◽  
Ion Mobility Mass Spectrometry ◽  
Conformational Heterogeneity ◽  
Wild Type ◽  
Intrinsically Disordered ◽  
Charged Residues ◽  
Oppositely Charged

The global dimensions and amplitudes of conformational fluctuations of intrinsically disordered proteins are governed, in part, by the linear segregation versus clustering of oppositely charged residues within the primary sequence. Ion Mobility-Mass Spectrometry (IM-MS) affords unique advantages for probing the conformational consequences of the linear patterning of oppositely charged residues because it measures and separates proteins electrosprayed from solution on the basis of charge and shape. Here, we use IM-MS to measure the conformational consequences of charge patterning on the C-terminal intrinsically disordered region (p27 IDR) of the cell cycle inhibitory protein p27<sup>Kip1</sup>. We report the range of charge states and accompanying collisional cross section distributions for wild-type p27 IDR and two variants with identical amino acid compositions, k14 and k56, distinguished by the extent of linear mixing versus segregation of oppositely charged residues. Wild-type p27 IDR (k31) and k14 where the oppositely charged residues are more evenly distributed, exhibit a broad distribution of charge states. This is concordant with high degrees of conformational heterogeneity in solution. By contrast, k56 with linear segregation of oppositely charged residues, leads to limited conformational heterogeneity and a narrow distribution of charged states. Molecular dynamics simulations demonstrate that the interplay between chain solvation and intra-chain interactions (self-solvation) leads to conformational distributions that are modulated by salt concentration, with the wild-type sequence showing the most sensitivity to changes in salt concentration. These results suggest that the charge patterning within the wild-type p27 IDR may be optimized to sample both highly solvated and self-solvated conformational states.

scholarly journals Ion Mobility Mass Spectrometry Uncovers the Impact of the Patterning of Oppositely Charged Residues on the Conformational Distributions of Intrinsically Disordered Proteins

2018 ◽  
Author(s):  
Rebecca Beveridge ◽  
Lukasz Migas ◽  
Rahul Das ◽  
Rohit Pappu ◽  
Richard Kriwacki ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Ion Mobility ◽  
Intrinsically Disordered Proteins ◽  
Disordered Proteins ◽  
Ion Mobility Mass Spectrometry ◽  
Conformational Heterogeneity ◽  
Wild Type ◽  
Intrinsically Disordered ◽  
Charged Residues ◽  
Oppositely Charged

The global dimensions and amplitudes of conformational fluctuations of intrinsically disordered proteins are governed, in part, by the linear segregation versus clustering of oppositely charged residues within the primary sequence. Ion Mobility-Mass Spectrometry (IM-MS) affords unique advantages for probing the conformational consequences of the linear patterning of oppositely charged residues because it measures and separates proteins electrosprayed from solution on the basis of charge and shape. Here, we use IM-MS to measure the conformational consequences of charge patterning on the C-terminal intrinsically disordered region (p27 IDR) of the cell cycle inhibitory protein p27<sup>Kip1</sup>. We report the range of charge states and accompanying collisional cross section distributions for wild-type p27 IDR and two variants with identical amino acid compositions, k14 and k56, distinguished by the extent of linear mixing versus segregation of oppositely charged residues. Wild-type p27 IDR (k31) and k14 where the oppositely charged residues are more evenly distributed, exhibit a broad distribution of charge states. This is concordant with high degrees of conformational heterogeneity in solution. By contrast, k56 with linear segregation of oppositely charged residues, leads to limited conformational heterogeneity and a narrow distribution of charged states. Molecular dynamics simulations demonstrate that the interplay between chain solvation and intra-chain interactions (self-solvation) leads to conformational distributions that are modulated by salt concentration, with the wild-type sequence showing the most sensitivity to changes in salt concentration. These results suggest that the charge patterning within the wild-type p27 IDR may be optimized to sample both highly solvated and self-solvated conformational states.

Rapid Diagnosis of Parkinson’s Disease from Sebum using Paper Spray Ionisation Ion Mobility Mass Spectrometry

2020 ◽  
Author(s):  
Depanjan Sarkar ◽  
Drupad Trivedi ◽  
Eleanor Sinclair ◽  
Sze Hway Lim ◽  
Caitlin Walton-Doyle ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Parkinson’S Disease ◽  
Parkinson's Disease ◽  
Ion Mobility ◽  
Neurodegenerative Disorder ◽  
Treatment Options ◽  
Ion Mobility Mass Spectrometry ◽  
Paper Spray ◽  
Molecular Features ◽  
Validation Set

Parkinson’s disease (PD) is the second most common neurodegenerative disorder for which identification of robust biomarkers to complement clinical PD diagnosis would accelerate treatment options and help to stratify disease progression. Here we demonstrate the use of paper spray ionisation coupled with ion mobility mass spectrometry (PSI IM-MS) to determine diagnostic molecular features of PD in sebum. PSI IM-MS was performed directly from skin swabs, collected from 34 people with PD and 30 matched control subjects as a training set and a further 91 samples from 5 different collection sites as a validation set. PSI IM-MS elucidates ~ 4200 features from each individual and we report two classes of lipids (namely phosphatidylcholine and cardiolipin) that differ significantly in the sebum of people with PD. Putative metabolite annotations are obtained using tandem mass spectrometry experiments combined with accurate mass measurements. Sample preparation and PSI IM-MS analysis and diagnosis can be performed ~5 minutes per sample offering a new route to for rapid and inexpensive confirmatory diagnosis of this disease.

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