This title appears in the Scientific Report :
2008
Please use the identifier:
http://hdl.handle.net/2128/15180 in citations.
Analyse des Substratspektums der ClpCP-Protease aus Corynebacterium glutamicum
Analyse des Substratspektums der ClpCP-Protease aus Corynebacterium glutamicum
Conditional degradation of proteins by ATP-dependent proteases is an important regulatory principe. Analyses on ClgR, the transcriptional regulator of genes involved in proteolysis and DNA repair, and GlnK, a P$_{II}$-type signal transduction regulatory protein of Nitrogen starvation, revealed that...
Saved in:
Personal Name(s): | Schweitzer, Jens-Eric (Corresponding author) |
---|---|
Contributing Institute: |
Biotechnologie 1; IBT-1 |
Imprint: |
Jülich
Forschungszentrum Jülich GmbH Zentralbibliothek, Verlag
2008
|
Physical Description: |
V, 130 S. |
Dissertation Note: |
Universität Düsseldorf, Diss., 2007 |
ISBN: |
978-3-89336-528-9 |
Document Type: |
Book Dissertation / PhD Thesis |
Research Program: |
Biotechnologie |
Series Title: |
Schriften des Forschungszentrums Jülich. Reihe Gesundheit / Health
8 |
Subject (ZB): | |
Link: |
OpenAccess |
Publikationsportal JuSER |
Conditional degradation of proteins by ATP-dependent proteases is an important regulatory principe. Analyses on ClgR, the transcriptional regulator of genes involved in proteolysis and DNA repair, and GlnK, a P$_{II}$-type signal transduction regulatory protein of Nitrogen starvation, revealed that these proteins are substrates of the $\textit{Corynebacterium glutamicum}$ ClpCP-protease. Aim of this work was to identify ffurther substrates of ClpCP and to determine how ClpC can recognise these substrates. The following results were achieved answering these questions: 1. Strains for the co-purification of substrates with inactive ClpP-derivates harbouring a mutation of the catalytic active serine to glycine and a C-terminal StrepTag-II were successfully constructed and named ClpCP1$^{TRAP}$. Step Tactin affinity chromatography of extracts of ClpCP1$^{TRAP}$ and ClpCP2$^{TRAP}$ led to the co-purification of 61 proteins, of which 14 occurred at least in five of ten purifications of inactive ClpP1 adn ClpP2. These putative substrates could be subdivided into different functional classes like translation, chaperones, amino acid metabolism and energy metabolism. Co-purification was also possible wih ClpP derivates by the C-terminal StrepTag-II. 2. Control experiments were made to support the specificity of the co-purification for ClpP1 and ClpP2. Purification of the plasmid encoded E1-subunit of the 2-oxoglutarate dehydrogenase complex (OdhA) with a C-Terminal StrepTag-II in the same genetic background as purification of proteins with a processed ClpP1 derivate containing an N-Terminal StrepTag-II was not possible, too. 3. Investigation on the termini of the co-purified proteins showed 10 N-terminal and 10 C-terminal putative recognition mitives for ClpC. Determination of the half-life of protein fusions with the 10 N-terminal sequence motives showed that eight motives had a negataive influence on the stability of the protein fusions. These motives might exhibit ClpC recognition sites. |