Aufgrund von Vorbereitungen auf eine neue Version von KOPS, können kommenden Montag und Dienstag keine Publikationen eingereicht werden. (Due to preparations for a new version of KOPS, no publications can be submitted next Monday and Tuesday.)
Type of Publication: | Journal article |
URI (citable link): | http://nbn-resolving.de/urn:nbn:de:bsz:352-234611 |
Author: | Schneider, Daniel; Schneider, Tatjana; Rösner, Daniel; Scheffner, Martin; Marx, Andreas |
Year of publication: | 2013 |
Published in: | Bioorganic & Medicinal Chemistry ; 21 (2013), 12. - pp. 3430-3435. - ISSN 0968-0896. - eISSN 1464-3391 |
Pubmed ID: | 23611767 |
DOI (citable link): | https://dx.doi.org/10.1016/j.bmc.2013.03.063 |
Summary: |
Posttranslational modification of proteins with ubiquitin (ubiquitylation) regulates numerous cellular processes. Besides functioning as a signal for proteasomal degradation, ubiquitylation has also non-proteolytic functions by altering the biochemical properties of the modified protein. To investigate the effect(s) of ubiquitylation on the properties of a protein, sufficient amounts of homogenously and well-defined ubiquitylated proteins are required. Here, we report on the elaboration of a method for the generation of high amounts of site-specifically mono-ubiquitylated proteins. Firstly, a one-step affinity purification scheme was developed for ubiquitin containing the unnatural amino acid azidohomoalanine at the C-terminal position. This ubiquitin was conjugated in a click reaction to recombinant DNA polymerase beta, equipped with an alkyne function at a distinct position. Secondly, addition of defined amounts of SDS to the reaction significantly improved product formation. With these two technical improvements, we have developed a straight forward procedure for the efficient generation of site-specifically ubiquitylated proteins that can be used to study the effect(s) of ubiquitylation on the activities/ properties of a protein.
|
Subject (DDC): | 540 Chemistry |
Keywords: | Ubiquitin, Unnatural amino acids, Click reaction, DNA polymerase beta, Protein conjugation |
Link to License: | In Copyright |
Bibliography of Konstanz: | Yes |
SCHNEIDER, Daniel, Tatjana SCHNEIDER, Daniel RÖSNER, Martin SCHEFFNER, Andreas MARX, 2013. Improving bioorthogonal protein ubiquitylation by click reaction. In: Bioorganic & Medicinal Chemistry. 21(12), pp. 3430-3435. ISSN 0968-0896. eISSN 1464-3391. Available under: doi: 10.1016/j.bmc.2013.03.063
@article{Schneider2013-06-15Impro-23461, title={Improving bioorthogonal protein ubiquitylation by click reaction}, year={2013}, doi={10.1016/j.bmc.2013.03.063}, number={12}, volume={21}, issn={0968-0896}, journal={Bioorganic & Medicinal Chemistry}, pages={3430--3435}, author={Schneider, Daniel and Schneider, Tatjana and Rösner, Daniel and Scheffner, Martin and Marx, Andreas} }
Schneider_234611.pdf | 888 |