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

Kód výsledku v IS VaVaI

<a href="https://www.isvavai.cz/riv?ss=detail&h=RIV%2F61388963%3A_____%2F22%3A00559233" target="_blank" >RIV/61388963:_____/22:00559233 - isvavai.cz</a>

Výsledek na webu

<a href="https://doi.org/10.1021/acs.analchem.2c00942" target="_blank" >https://doi.org/10.1021/acs.analchem.2c00942</a>

DOI - Digital Object Identifier

<a href="http://dx.doi.org/10.1021/acs.analchem.2c00942" target="_blank" >10.1021/acs.analchem.2c00942</a>

Jazyk výsledku

angličtina

Název v původním jazyce

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

Popis výsledku v původním jazyce

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-IM-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. Gas-phase HDX with ECD fragmentation for exchange site localization 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 many regions with protection from HDX, and the 11(+) charge state, ionized from conditions that stabilize the A-state, showed high levels of deuterium incorporation throughout most of the protein sequence. The 7(+) ions of cytochrome c ionized from aqueous conditions showed greater HDX with unstructured regions of the protein relative to interior, structured regions, especially those involved in heme binding. With careful tuning and attention to deuterium scrambling, our approach holds promise for determining region-specific information on a conformer-selected basis for gas-phase protein structures, including localized characterizations of ligand, epitope, and protein-protein binding.

Název v anglickém jazyce

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

Popis výsledku anglicky

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-IM-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. Gas-phase HDX with ECD fragmentation for exchange site localization 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 many regions with protection from HDX, and the 11(+) charge state, ionized from conditions that stabilize the A-state, showed high levels of deuterium incorporation throughout most of the protein sequence. The 7(+) ions of cytochrome c ionized from aqueous conditions showed greater HDX with unstructured regions of the protein relative to interior, structured regions, especially those involved in heme binding. With careful tuning and attention to deuterium scrambling, our approach holds promise for determining region-specific information on a conformer-selected basis for gas-phase protein structures, including localized characterizations of ligand, epitope, and protein-protein binding.

Druh

J_imp - Článek v periodiku v databázi Web of Science

CEP obor

—

OECD FORD obor

10406 - Analytical chemistry

Projekt

—

Návaznosti

I - Institucionalni podpora na dlouhodoby koncepcni rozvoj vyzkumne organizace

Rok uplatnění

2022

Kód důvěrnosti údajů

S - Úplné a pravdivé údaje o projektu nepodléhají ochraně podle zvláštních právních předpisů

Název periodika

Analytical Chemistry

ISSN

0003-2700

e-ISSN

1520-6882

Svazek periodika

94

Číslo periodika v rámci svazku

25

Stát vydavatele periodika

US - Spojené státy americké

Počet stran výsledku

8

Strana od-do

8975-8982

Kód UT WoS článku

000820693400001

EID výsledku v databázi Scopus

2-s2.0-85133959392