Isotopic Depletion Increases the Spatial Resolution of FPOP Top-Down Mass Spectrometry Analysis
The result's identifiers
Result code in IS VaVaI
<a href="https://www.isvavai.cz/riv?ss=detail&h=RIV%2F61388971%3A_____%2F24%3A00585509" target="_blank" >RIV/61388971:_____/24:00585509 - isvavai.cz</a>
Alternative codes found
RIV/86652036:_____/24:00585509 RIV/00216208:11310/24:10477718
Result on the web
<a href="https://pubs.acs.org/doi/10.1021/acs.analchem.3c03759" target="_blank" >https://pubs.acs.org/doi/10.1021/acs.analchem.3c03759</a>
DOI - Digital Object Identifier
<a href="http://dx.doi.org/10.1021/acs.analchem.3c03759" target="_blank" >10.1021/acs.analchem.3c03759</a>
Alternative languages
Result language
angličtina
Original language name
Isotopic Depletion Increases the Spatial Resolution of FPOP Top-Down Mass Spectrometry Analysis
Original language description
Protein radical labeling, like fast photochemical oxidation of proteins (FPOP), coupled to a top-down mass spectrometry (MS) analysis offers an alternative analytical method for probing protein structure or protein interaction with other biomolecules, for instance, proteins and DNA. However, with the increasing mass of studied analytes, the MS/MS spectra become complex and exhibit a low signal-to-noise ratio. Nevertheless, these difficulties may be overcome by protein isotope depletion. Thus, we aimed to use protein isotope depletion to analyze FPOP-oxidized samples by top-down MS analysis. For this purpose, we prepared isotopically natural (IN) and depleted (ID) forms of the FOXO4 DNA binding domain (FOXO4-DBD) and studied the protein-DNA interaction interface with double-stranded DNA, the insulin response element (IRE), after exposing the complex to hydroxyl radicals. As shown by comparing tandem mass spectra of natural and depleted proteins, the ID form increased the signal-to-noise ratio of useful fragment ions, thereby enhancing the sequence coverage by more than 19%. This improvement in the detection of fragment ions enabled us to detect 22 more oxidized residues in the ID samples than in the IN sample. Moreover, less common modifications were detected in the ID sample, including the formation of ketones and lysine carbonylation. Given the higher quality of ID top-down MSMS data set, these results provide more detailed information on the complex formation between transcription factors and DNA-response elements. Therefore, our study highlights the benefits of isotopic depletion for quantitative top-down proteomics. Data are available via ProteomeXchange with the identifier PXD044447.
Czech name
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Czech description
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Classification
Type
J<sub>imp</sub> - Article in a specialist periodical, which is included in the Web of Science database
CEP classification
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OECD FORD branch
10606 - Microbiology
Result continuities
Project
Result was created during the realization of more than one project. More information in the Projects tab.
Continuities
I - Institucionalni podpora na dlouhodoby koncepcni rozvoj vyzkumne organizace
Others
Publication year
2024
Confidentiality
S - Úplné a pravdivé údaje o projektu nepodléhají ochraně podle zvláštních právních předpisů
Data specific for result type
Name of the periodical
Analytical Chemistry
ISSN
0003-2700
e-ISSN
1520-6882
Volume of the periodical
96
Issue of the periodical within the volume
4
Country of publishing house
US - UNITED STATES
Number of pages
10
Pages from-to
1478-1487
UT code for WoS article
001154899300001
EID of the result in the Scopus database
2-s2.0-85183043291