Efficient and robust preparation of tyrosine phosphorylated   
intrinsically disordered proteins

J 2019

Efficient and robust preparation of tyrosine phosphorylated intrinsically disordered proteins

BRÁZDA, Pavel, Ondrej ŠEDO, Karel KUBÍČEK a Richard ŠTEFL

Základní údaje

Originální název

Efficient and robust preparation of tyrosine phosphorylated intrinsically disordered proteins

Autoři

BRÁZDA, Pavel (203 Česká republika, domácí), Ondrej ŠEDO (203 Česká republika, domácí), Karel KUBÍČEK (203 Česká republika, domácí) a Richard ŠTEFL (203 Česká republika, garant, domácí)

Vydání

BioTechniques, UNITED HOUSE, 2 ALBERT PL, LONDON N3 1QB, FUTURE SCI LTD, 2019, 0736-6205

Další údaje

Jazyk

angličtina

Typ výsledku

Článek v odborném periodiku

Obor

10608 Biochemistry and molecular biology

Stát vydavatele

Velká Británie a Severní Irsko

Utajení

není předmětem státního či obchodního tajemství

Odkazy

URL

Impakt faktor

Impact factor: 1.541

Kód RIV

RIV/00216224:14740/19:00107820

Organizační jednotka

Středoevropský technologický institut

DOI

http://dx.doi.org/10.2144/btn-2019-0033

UT WoS

000475967900005

Klíčová slova anglicky

C-terminal domain; co-expression; CTD; IDP; intrinsically disordered proteins; phosphorylation; purification; RNA polymerase II; TRANSCRIPTION TERMINATION; LARGEST SUBUNIT; CELL-CYCLE; KINASE; STRATEGIES; SEMISYNTHESIS

Štítky

CF NMR, CF PROT, rivok

Příznaky

Mezinárodní význam, Recenzováno

Změněno: 14. 10. 2024 13:09, Ing. Jana Kuchtová

Anotace

V originále

Intrinsically disordered proteins (IDPs) are subject to post-translational modifications. This allows the same polypeptide to undertake different interaction networks with different consequences, ranging from regulatory signalling networks to formation of membraneless organelles. We report a robust method for co-expression of modification enzyme and SUMO-tagged IDP with subsequent purification procedure allowing production of modified IDP. The robustness of our protocol is demonstrated on a challenging system, RNA polymerase II C-terminal domain (CM), that is a low-complexity repetitive region with multiple phosphorylation sites. In vitro phosphorylation approaches fail to yield multiple-site phosphorylated CTD, whereas our in vivo protocol allows to rapidly produce near homogeneous phosphorylated CTD at a low cost. These samples can be used in functional and structural studies.

Návaznosti

GA15-17670S, projekt VaV

Název: Molekulární podstata pro specifické rozpoznávání histonu H3K4me2 pomocí Set3 PHD domény

Investor: Grantová agentura ČR, Molekulární podstata pro specifické rozpoznávání histonu H3K4me2 pomocí Set3 PHD domény

GA18-11397S, projekt VaV

Název: Molekulární achitektura komplexů ADAR - RNA

Investor: Grantová agentura ČR, Molecular architecture of ADAR - RNA complexes

LQ1601, projekt VaV

Název: CEITEC 2020 (Akronym: CEITEC2020)

Investor: Ministerstvo školství, mládeže a tělovýchovy ČR, CEITEC 2020

649030, interní kód MU

Název: DECOR - Dynamic assembly and exchange of RNA polymerase II CTD factors (Akronym: DECOR)

Investor: Evropská unie, DECOR - Dynamic assembly and exchange of RNA polymerase II CTD factors, ERC (Excellent Science)

90043, velká výzkumná infrastruktura

Název: CIISB

Přiložené soubory

btn-2019-0033.pdf

Vyhledat podobné dokumenty

Citovat

BRÁZDA, Pavel, Ondrej ŠEDO, Karel KUBÍČEK a Richard ŠTEFL. Efficient and robust preparation of tyrosine phosphorylated intrinsically disordered proteins. BioTechniques. UNITED HOUSE, 2 ALBERT PL, LONDON N3 1QB: FUTURE SCI LTD, 2019, roč. 67, č. 1, s. 16-22. ISSN 0736-6205. Dostupné z: https://dx.doi.org/10.2144/btn-2019-0033.

@article{1585263,
   author = {Brázda, Pavel and Šedo, Ondrej and Kubíček, Karel and Štefl, Richard},
   article_location = {UNITED HOUSE, 2 ALBERT PL, LONDON N3 1QB},
   article_number = {1},
   doi = {http://dx.doi.org/10.2144/btn-2019-0033},
   keywords = {C-terminal domain; co-expression; CTD; IDP; intrinsically disordered proteins; phosphorylation; purification; RNA polymerase II; TRANSCRIPTION TERMINATION; LARGEST SUBUNIT; CELL-CYCLE; KINASE; STRATEGIES; SEMISYNTHESIS},
   language = {eng},
   issn = {0736-6205},
   journal = {BioTechniques},
   title = {Efficient and robust preparation of tyrosine phosphorylated intrinsically disordered proteins},
   url = {https://www.future-science.com/doi/full/10.2144/btn-2019-0033},
   volume = {67},
   year = {2019}
}

TY  - JOUR
ID  - 1585263
AU  - Brázda, Pavel - Šedo, Ondrej - Kubíček, Karel - Štefl, Richard
PY  - 2019
TI  - Efficient and robust preparation of tyrosine phosphorylated intrinsically disordered proteins
JF  - BioTechniques
VL  - 67
IS  - 1
SP  - 16-22
EP  - 16-22
PB  - FUTURE SCI LTD
SN  - 07366205
KW  - C-terminal domain
KW  - co-expression
KW  - CTD
KW  - IDP
KW  - intrinsically disordered proteins
KW  - phosphorylation
KW  - purification
KW  - RNA polymerase II
KW  - TRANSCRIPTION TERMINATION
KW  - LARGEST SUBUNIT
KW  - CELL-CYCLE
KW  - KINASE
KW  - STRATEGIES
KW  - SEMISYNTHESIS
UR  - https://www.future-science.com/doi/full/10.2144/btn-2019-0033
L2  - https://www.future-science.com/doi/full/10.2144/btn-2019-0033
N2  - Intrinsically disordered proteins (IDPs) are subject to post-translational modifications. This allows the same polypeptide to undertake different interaction networks with different consequences, ranging from regulatory signalling networks to formation of membraneless organelles. We report a robust method for co-expression of modification enzyme and SUMO-tagged IDP with subsequent purification procedure allowing production of modified IDP. The robustness of our protocol is demonstrated on a challenging system, RNA polymerase II C-terminal domain (CM), that is a low-complexity repetitive region with multiple phosphorylation sites. In vitro phosphorylation approaches fail to yield multiple-site phosphorylated CTD, whereas our in vivo protocol allows to rapidly produce near homogeneous phosphorylated CTD at a low cost. These samples can be used in functional and structural studies.
ER  -

BRÁZDA, Pavel, Ondrej ŠEDO, Karel KUBÍČEK a Richard ŠTEFL. Efficient and robust preparation of tyrosine phosphorylated intrinsically disordered proteins. \textit{BioTechniques}. UNITED HOUSE, 2 ALBERT PL, LONDON N3 1QB: FUTURE SCI LTD, 2019, roč.~67, č.~1, s.~16-22. ISSN~0736-6205. Dostupné z: https://dx.doi.org/10.2144/btn-2019-0033.

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