Abstract:
:Intrinsically disordered proteins (IDPs) are subject to post-translational modifications. This allows the same polypeptide to be involved in different interaction networks with different consequences, ranging from regulatory signalling networks to the formation of membrane-less organelles. We report a robust method for co-expression of modification enzyme and SUMO-tagged IDPs with a subsequent purification procedure that allows for the production of modified IDP. The robustness of our protocol is demonstrated using a challenging system: RNA polymerase II C-terminal domain (CTD); 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 the rapid production of near homogeneous phosphorylated CTD at a low cost. These samples can be used in functional and structural studies.
journal_name
Biotechniquesjournal_title
BioTechniquesauthors
Brázda P,Šedo O,Kubíček K,Štefl Rdoi
10.2144/btn-2019-0033subject
Has Abstractpub_date
2019-07-01 00:00:00pages
16-22issue
1eissn
0736-6205issn
1940-9818journal_volume
67pub_type
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