Electrochemical detection of DNA binding by tumor suppressor
p53 protein using osmium-labeled oligonucleotide probes and
catalytic hydrogen evolution at the mercury electrode

J 2014

Electrochemical detection of DNA binding by tumor suppressor p53 protein using osmium-labeled oligonucleotide probes and catalytic hydrogen evolution at the mercury electrode

NĚMCOVÁ, Kateřina, Peter ŠEBEST, Luděk HAVRAN, Petr ORSÁG, Miroslav FOJTA et. al.

Základní údaje

Originální název

Electrochemical detection of DNA binding by tumor suppressor p53 protein using osmium-labeled oligonucleotide probes and catalytic hydrogen evolution at the mercury electrode

Autoři

NĚMCOVÁ, Kateřina (203 Česká republika), Peter ŠEBEST (203 Česká republika), Luděk HAVRAN (203 Česká republika), Petr ORSÁG (203 Česká republika, domácí), Miroslav FOJTA (203 Česká republika, garant, domácí) a Hana PIVOŇKOVÁ (203 Česká republika)

Vydání

Analytical and Bioanalytical chemistry, HEIDELBERG, SPRINGER HEIDELBERG, 2014, 1618-2642

Další údaje

Jazyk

angličtina

Typ výsledku

Článek v odborném periodiku

Obor

10405 Electrochemistry

Stát vydavatele

Německo

Utajení

není předmětem státního či obchodního tajemství

Odkazy

URL

Impakt faktor

Impact factor: 3.436

Kód RIV

RIV/00216224:14740/14:00079183

Organizační jednotka

Středoevropský technologický institut

DOI

http://dx.doi.org/10.1007/s00216-014-7996-0

UT WoS

000341834500011

Klíčová slova anglicky

Electrochemical analysis; Labeled probes; Osmium complex; Tumor suppressor protein p53; DNA-protein interaction; Immunoprecipitation; Competition assay; Magnetic beads; Mercury electrode

Štítky

kontrola MP, MP, rivok

Příznaky

Mezinárodní význam, Recenzováno

Změněno: 23. 2. 2015 13:18, Martina Prášilová

Anotace

V originále

In this paper, we present an electrochemical DNA-protein interaction assay based on a combination of protein-specific immunoprecipitation at magnetic beads (MBIP) with application of oligonucleotide (ON) probes labeled with an electroactive oxoosmium complex (Os,bipy). We show that double-stranded ONs bearing a dT(20) tail labeled with Os,bipy are specifically recognized by the tumor suppressor p53 protein according to the presence or absence of a specific binding site (p53CON) in the double-stranded segment. We demonstrate the applicability of the Os,bipy-labeled probes in titration as well as competition MBIP assays to evaluate p53 relative affinity to various sequence-specific or structurally distinct unlabeled DNA substrates upon modulation of the p53-DNA binding by monoclonal antibodies used for the immunoprecipitation. To detect the p53-bound osmium-labeled probes, we took advantage of a catalytic peak yielded by Os,bipy-modified DNA at the mercury-based electrodes, allowing facile determination of subnanogram quantities of the labeled oligonucleotides. Versatility of the electrochemical MBIP technique and its general applicability in studies of any DNA-binding protein is discussed.

Citovat

NĚMCOVÁ, Kateřina, Peter ŠEBEST, Luděk HAVRAN, Petr ORSÁG, Miroslav FOJTA a Hana PIVOŇKOVÁ. Electrochemical detection of DNA binding by tumor suppressor p53 protein using osmium-labeled oligonucleotide probes and catalytic hydrogen evolution at the mercury electrode. Analytical and Bioanalytical chemistry. HEIDELBERG: SPRINGER HEIDELBERG, 2014, roč. 406, č. 24, s. 5843-5852. ISSN 1618-2642. Dostupné z: https://dx.doi.org/10.1007/s00216-014-7996-0.

@article{1227600,
   author = {Němcová, Kateřina and Šebest, Peter and Havran, Luděk and Orság, Petr and Fojta, Miroslav and Pivoňková, Hana},
   article_location = {HEIDELBERG},
   article_number = {24},
   doi = {http://dx.doi.org/10.1007/s00216-014-7996-0},
   keywords = {Electrochemical analysis; Labeled probes; Osmium complex; Tumor suppressor protein p53; DNA-protein interaction; Immunoprecipitation; Competition assay; Magnetic beads; Mercury electrode},
   language = {eng},
   issn = {1618-2642},
   journal = {Analytical and Bioanalytical chemistry},
   title = {Electrochemical detection of DNA binding by tumor suppressor p53 protein using osmium-labeled oligonucleotide probes and catalytic hydrogen evolution at the mercury electrode},
   url = {http://download.springer.com/static/pdf/605/art%253A10.1007%252Fs00216-014-7996-0.pdf?auth66=1424693573_e49fe0b8548f527d28393c19da86150b&ext=.pdf},
   volume = {406},
   year = {2014}
}

TY  - JOUR
ID  - 1227600
AU  - Němcová, Kateřina - Šebest, Peter - Havran, Luděk - Orság, Petr - Fojta, Miroslav - Pivoňková, Hana
PY  - 2014
TI  - Electrochemical detection of DNA binding by tumor suppressor p53 protein using osmium-labeled oligonucleotide probes and catalytic hydrogen evolution at the mercury electrode
JF  - Analytical and Bioanalytical chemistry
VL  - 406
IS  - 24
SP  - 5843-5852
EP  - 5843-5852
PB  - SPRINGER HEIDELBERG
SN  - 16182642
KW  - Electrochemical analysis
KW  - Labeled probes
KW  - Osmium complex
KW  - Tumor suppressor protein p53
KW  - DNA-protein interaction
KW  - Immunoprecipitation
KW  - Competition assay
KW  - Magnetic beads
KW  - Mercury electrode
UR  - http://download.springer.com/static/pdf/605/art%253A10.1007%252Fs00216-014-7996-0.pdf?auth66=1424693573_e49fe0b8548f527d28393c19da86150b&ext=.pdf
L2  - http://download.springer.com/static/pdf/605/art%253A10.1007%252Fs00216-014-7996-0.pdf?auth66=1424693573_e49fe0b8548f527d28393c19da86150b&ext=.pdf
N2  - In this paper, we present an electrochemical DNA-protein interaction assay based on a combination of protein-specific immunoprecipitation at magnetic beads (MBIP) with application of oligonucleotide (ON) probes labeled with an electroactive oxoosmium complex (Os,bipy). We show that double-stranded ONs bearing a dT(20) tail labeled with Os,bipy are specifically recognized by the tumor suppressor p53 protein according to the presence or absence of a specific binding site (p53CON) in the double-stranded segment. We demonstrate the applicability of the Os,bipy-labeled probes in titration as well as competition MBIP assays to evaluate p53 relative affinity to various sequence-specific or structurally distinct unlabeled DNA substrates upon modulation of the p53-DNA binding by monoclonal antibodies used for the immunoprecipitation. To detect the p53-bound osmium-labeled probes, we took advantage of a catalytic peak yielded by Os,bipy-modified DNA at the mercury-based electrodes, allowing facile determination of subnanogram quantities of the labeled oligonucleotides. Versatility of the electrochemical MBIP technique and its general applicability in studies of any DNA-binding protein is discussed.
ER  -

NĚMCOVÁ, Kateřina, Peter ŠEBEST, Luděk HAVRAN, Petr ORSÁG, Miroslav FOJTA a Hana PIVOŇKOVÁ. Electrochemical detection of DNA binding by tumor suppressor p53 protein using osmium-labeled oligonucleotide probes and catalytic hydrogen evolution at the mercury electrode. \textit{Analytical and Bioanalytical chemistry}. HEIDELBERG: SPRINGER HEIDELBERG, 2014, roč.~406, č.~24, s.~5843-5852. ISSN~1618-2642. Dostupné z: https://dx.doi.org/10.1007/s00216-014-7996-0.

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