Ion Mobility and Surface Collisions: Submicrometer Capillaries Can Produce Native-like Protein Complexes

Panczyk, Erin; Gilbert, Joshua D.; Jagdale, Gargi S.; Stiving, Alyssa Q.; Baker, Lane A.; Wysocki, Vicki H.

doi:10.1021/acs.analchem.9b03666

Cited by 13 publications

(24 citation statements)

References 39 publications

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“…25,45 Nonspecific clustering can be reduced by using narrow emitters that produce smaller initial droplets, thereby decreasing the number of analyte molecules in each droplet. 24,62,66,67,70 Unfortunately, narrow emitters are prone to clogging. 72 Similarly, one can lower the analyte concentration, such that the fraction of droplets containing more than one analyte molecule is lowered.…”

Section: ■ Introductionmentioning

confidence: 99%

“…Mass analysis of these ions reveals their subunit stoichiometry. − Native ESI-MS is attractive because of its conceptual simplicity, minimal sample preparation, high sensitivity, and short analysis time. Additional insights are obtainable by combining ESI-MS with ion mobility spectrometry (IMS), − gas-phase activation, − and fragmentation techniques. − …”

Section: Introductionmentioning

confidence: 99%

“…This phenomenon can manifest itself as complex formation from monomeric proteins − or the assembly of complexes into higher-order oligomers. ,− There can also be a mix of specific solution binding and nonspecific clustering. , All of these clustering scenarios complicate the interpretation of native ESI-MS data. Various strategies have been proposed for mitigating this problem, ,,− but it is nonetheless challenging to distinguish specific from nonspecific complexes in a mass spectrum. Interestingly, there are also instances where nonspecific clustering is beneficial; for example, protein clusters can serve as model systems for benchmarking mass analyzer performance at high m / z and as a testbed for top-down dissociation experiments. ,,, …”

Section: Introductionmentioning

confidence: 99%

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Atomistic Insights into the Formation of Nonspecific Protein Complexes during Electrospray Ionization

Aliyari

Konermann

2021

Anal. Chem.

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Native electrospray ionization (ESI)-mass spectrometry (MS) is widely used for the detection and characterization of multi-protein complexes. A well-known problem with this approach is the possible occurrence of nonspecific protein clustering in the ESI plume. This effect can distort the results of binding affinity measurements, and it can even generate gas-phase complexes from proteins that are strictly monomeric in bulk solution. By combining experiments and molecular dynamics (MD) simulations, the current work for the first time provides detailed insights into the ESI clustering of proteins. Using ubiquitin as a model system, we demonstrate how the entrapment of more than one protein molecule in an ESI droplet can generate nonspecific clusters (e.g., dimers or trimers) via solvent evaporation to dryness. These events are in line with earlier proposals, according to which protein clustering is associated with the charged residue model (CRM). MD simulations on cytochrome c (which carries a large intrinsic positive charge) confirmed the viability of this CRM avenue. In addition, the cytochrome c data uncovered an alternative mechanism where protein–protein contacts were formed early within ESI droplets, followed by cluster ejection from the droplet surface. This second pathway is consistent with the ion evaporation model (IEM). The observation of these IEM events for large protein clusters is unexpected because the IEM has been thought to be associated primarily with low-molecular-weight analytes. In all cases, our MD simulations produced protein clusters that were stabilized by intermolecular salt bridges. The MD-generated charge states agreed with experiments. Overall, this work reveals that ESI-induced protein clustering does not follow a tightly orchestrated pathway but can proceed along different avenues.

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Section: ■ Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Atomistic Insights into the Formation of Nonspecific Protein Complexes during Electrospray Ionization

Aliyari

Konermann

2021

Anal. Chem.

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“…The smaller initial charged droplets produced by the submicron emitter tips effectively reduce nonspecific adducts and salt clusters, 10,11 which is critical for the analysis of native biomolecules and complexes. 5,12–14 However, except for the 26 pL min −1 flow rate for a 600 nm emitter tip, 15 the flow rates for most submicron emitter tips are unknown. Femtoamp electrospray ionization (fA ESI) was developed using both micrometer and submicron emitter tips.…”

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confidence: 99%

Conducting and characterizing femto flow electrospray ionization

Allen

et al. 2022

Analyst

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“…The smaller initial charged droplets produced by the submicron emitter tips effectively reduce nonspecific adducts and salt clusters, 10,11 which is critical for the analysis of native biomolecules and complexes. 5,[12][13][14] However, except for the 26 pL/min for a 600 nm emitter tip, 15 the flow rates for most submicron emitter tips are unknown. Femtoamp electrospray ionization (fA ESI) was developed using both micrometer and submicron emitter tips.…”

mentioning

confidence: 99%