Pull-down Assay on Streptavidin Beads and Surface Plasmon Resonance Chips for SWATH-MS-based Interactomics
Identifikátory výsledku
Kód výsledku v IS VaVaI
<a href="https://www.isvavai.cz/riv?ss=detail&h=RIV%2F00209805%3A_____%2F18%3A00078032" target="_blank" >RIV/00209805:_____/18:00078032 - isvavai.cz</a>
Nalezeny alternativní kódy
RIV/00216224:14310/18:00101543
Výsledek na webu
<a href="http://cgp.iiarjournals.org/content/15/5/395.long" target="_blank" >http://cgp.iiarjournals.org/content/15/5/395.long</a>
DOI - Digital Object Identifier
<a href="http://dx.doi.org/10.21873/cgp.20098" target="_blank" >10.21873/cgp.20098</a>
Alternativní jazyky
Jazyk výsledku
angličtina
Název v původním jazyce
Pull-down Assay on Streptavidin Beads and Surface Plasmon Resonance Chips for SWATH-MS-based Interactomics
Popis výsledku v původním jazyce
Pul-down assay is a popular in vitro method for identification of physical interactors of selected proteins. Here, for the first time, we compared three conventional variants of pull-down assay with the streptavidin-modified surface plasmon resonance (SPR) chips for the detection of PDZ and LIM domain protein 2 (PDLIM2) interaction partners. PDLIM2 protein-protein interactions were analysed by three variants of pull-down assay on streptavidin beads using LC-MS/MS in "Sequential Window Acquisition of all Theoretical fragment ion spectra (SWATH)" mode and compared with LC-SWATH-MS/MS data from SPR chips. The results showed that (i) the use of SPR chip led to comparable data compared to on-column streptavidin beads, (ii) gravity flow and microflow in wash and elution steps provided better results than centrifugation, and (iii) type and concentration of detergent did not significantly affect the interactome data of cancer-associated PDLIM2. Our study supports further application of SPR-based affinity purification with SWATH mass spectrometry for reproducible and controlled characterization of cancer-associated interactomes.
Název v anglickém jazyce
Pull-down Assay on Streptavidin Beads and Surface Plasmon Resonance Chips for SWATH-MS-based Interactomics
Popis výsledku anglicky
Pul-down assay is a popular in vitro method for identification of physical interactors of selected proteins. Here, for the first time, we compared three conventional variants of pull-down assay with the streptavidin-modified surface plasmon resonance (SPR) chips for the detection of PDZ and LIM domain protein 2 (PDLIM2) interaction partners. PDLIM2 protein-protein interactions were analysed by three variants of pull-down assay on streptavidin beads using LC-MS/MS in "Sequential Window Acquisition of all Theoretical fragment ion spectra (SWATH)" mode and compared with LC-SWATH-MS/MS data from SPR chips. The results showed that (i) the use of SPR chip led to comparable data compared to on-column streptavidin beads, (ii) gravity flow and microflow in wash and elution steps provided better results than centrifugation, and (iii) type and concentration of detergent did not significantly affect the interactome data of cancer-associated PDLIM2. Our study supports further application of SPR-based affinity purification with SWATH mass spectrometry for reproducible and controlled characterization of cancer-associated interactomes.
Klasifikace
Druh
J<sub>imp</sub> - Článek v periodiku v databázi Web of Science
CEP obor
—
OECD FORD obor
10608 - Biochemistry and molecular biology
Návaznosti výsledku
Projekt
Výsledek vznikl pri realizaci vícero projektů. Více informací v záložce Projekty.
Návaznosti
P - Projekt vyzkumu a vyvoje financovany z verejnych zdroju (s odkazem do CEP)
Ostatní
Rok uplatnění
2018
Kód důvěrnosti údajů
S - Úplné a pravdivé údaje o projektu nepodléhají ochraně podle zvláštních právních předpisů
Údaje specifické pro druh výsledku
Název periodika
Cancer genomics & proteomics
ISSN
1109-6535
e-ISSN
—
Svazek periodika
15
Číslo periodika v rámci svazku
5
Stát vydavatele periodika
GR - Řecká republika
Počet stran výsledku
10
Strana od-do
395-404
Kód UT WoS článku
000444004800005
EID výsledku v databázi Scopus
2-s2.0-85052992178