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. |
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@article{38921, 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}, 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 - 38921 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 - 0736-6205 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 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.
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