Utilization of Fast Photochemical Oxidation of Proteins and Both Bottom-up and Top-down Mass Spectrometry for Structural Characterization of a Transcription Factor-dsDNA Complex

Kód výsledku v IS VaVaI

<a href="https://www.isvavai.cz/riv?ss=detail&h=RIV%2F00216224%3A90127%2F22%3A00140007" target="_blank" >RIV/00216224:90127/22:00140007 - isvavai.cz</a>

Nalezeny alternativní kódy

RIV/61388971:_____/22:00556153 RIV/00216208:11310/22:10445307

Výsledek na webu

<a href="https://pubs.acs.org/doi/10.1021/acs.analchem.1c04746" target="_blank" >https://pubs.acs.org/doi/10.1021/acs.analchem.1c04746</a>

DOI - Digital Object Identifier

<a href="http://dx.doi.org/10.1021/acs.analchem.1c04746" target="_blank" >10.1021/acs.analchem.1c04746</a>

Jazyk výsledku

angličtina

Název v původním jazyce

Utilization of Fast Photochemical Oxidation of Proteins and Both Bottom-up and Top-down Mass Spectrometry for Structural Characterization of a Transcription Factor-dsDNA Complex

Popis výsledku v původním jazyce

A combination of covalent labeling techniques and mass spectrometry (MS) is currently a progressive approach for deriving insights related to the mapping of protein surfaces or protein-ligand interactions. In this study, we mapped an interaction interface between the DNA binding domain (DBD) of FOXO4 protein and the DNA binding element (DAF16) using fast photochemical oxidation of proteins (FPOP). Residues involved in protein-DNA interaction were identified using the bottom-up approach. To confirm the findings and avoid a misinterpretation of the obtained data, caused by possible multiple radical oxidations leading to the protein surface alteration and oxidation of deeply buried amino acid residues, a top-down approach was employed for the first time in FPOP analysis. An isolation of singly oxidized ions enabled their gas-phase separation from multiply oxidized species followed by CID and ECD fragmentation. Application of both fragmentation techniques allowed generation of complementary fragment sets, out of which the regions shielded in the presence of DNA were deduced. The findings obtained by bottom-up and top-down approaches were highly consistent. Finally, FPOP results were compared with those of the HDX study of the FOXO4-DBD center dot DAF16 complex. No contradictions were found between the methods. Moreover, their combination provides complementary information related to the structure and dynamics of the protein-DNA complex. Data are available via ProteomeXchange with identifier PXD027624.

Název v anglickém jazyce

Utilization of Fast Photochemical Oxidation of Proteins and Both Bottom-up and Top-down Mass Spectrometry for Structural Characterization of a Transcription Factor-dsDNA Complex

Popis výsledku anglicky

A combination of covalent labeling techniques and mass spectrometry (MS) is currently a progressive approach for deriving insights related to the mapping of protein surfaces or protein-ligand interactions. In this study, we mapped an interaction interface between the DNA binding domain (DBD) of FOXO4 protein and the DNA binding element (DAF16) using fast photochemical oxidation of proteins (FPOP). Residues involved in protein-DNA interaction were identified using the bottom-up approach. To confirm the findings and avoid a misinterpretation of the obtained data, caused by possible multiple radical oxidations leading to the protein surface alteration and oxidation of deeply buried amino acid residues, a top-down approach was employed for the first time in FPOP analysis. An isolation of singly oxidized ions enabled their gas-phase separation from multiply oxidized species followed by CID and ECD fragmentation. Application of both fragmentation techniques allowed generation of complementary fragment sets, out of which the regions shielded in the presence of DNA were deduced. The findings obtained by bottom-up and top-down approaches were highly consistent. Finally, FPOP results were compared with those of the HDX study of the FOXO4-DBD center dot DAF16 complex. No contradictions were found between the methods. Moreover, their combination provides complementary information related to the structure and dynamics of the protein-DNA complex. Data are available via ProteomeXchange with identifier PXD027624.

Druh

J_imp - Článek v periodiku v databázi Web of Science

CEP obor

—

OECD FORD obor

10406 - Analytical chemistry

Projekt

Výsledek vznikl pri realizaci vícero projektů. Více informací v záložce Projekty.

Návaznosti

—

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

—

Svazek periodika

94

Číslo periodika v rámci svazku

7

Stát vydavatele periodika

US - Spojené státy americké

Počet stran výsledku

8

Strana od-do

3203-3210

Kód UT WoS článku

000758042700001

EID výsledku v databázi Scopus

2-s2.0-85124911406