Hofweber, Roland and Horn, Gudrun and Langmann, Thomas and Balbach, Jochen and Kremer, Werner and Schmitz, Gerd and Kalbitzer, Hans-Robert (2005) The influence of cold shock proteins on transcription and translation studied in cell-free model systems. FEBS Journal 272 (18), pp. 4691-4702.
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Cold shock proteins (CSPs) form a family of highly conserved bacterial proteins capable of single-stranded nucleic acid binding. They are suggested to act as RNA chaperones during cold shock inhibiting the formation of RNA secondary structures, which are unfavourable for transcription and translation. To test this commonly accepted theory, isolated CSPs from a mesophilic, thermophilic and a hyperthermophilic bacterium (Bacillus subtilis, Bacillus caldolyticus and Thermotoga maritima) were studied in an Escherichia coli based cell free expression system on their capability of enhancing protein expression by reduction of mRNA secondary structures. The E. coli based expression of chloramphenicol acetyltransferase and of H-Ras served as model systems. We observed a concentration-dependent suppression of transcription and translation by the different CSPs which makes the considered addition of CSPs for enhancing the protein expression in in vitro translation systems obsolete. Protein expression was completely inhibited at CSP concentrations present under cold shock conditions. The CSP concentrations necessary for 50% inhibition were lowest (140 µm) for the protein of the hyperthermophilic and increased when the thermophilic (215 µm) or even the mesophilic protein (451 µm) was used. Isolated in vitro transcription under the influence of CSPs showed that the transcriptory effect is independent from the rest of the cell. It could be shown in a control experiment that the inhibition of protein expression can be removed by addition of hepta-2'-desoxy-thymidylate (dT7); a heptanucleotide that competitively binds to CSP. The data are in line with a hypothesis that CSPs act on bulk protein expression not as RNA chaperones but inhibit their transcription and translation by rather unspecific nucleic acid binding.
|Institutions:||Biology, Preclinical Medicine > Institut für Biophysik und physikalische Biochemie > Prof. Dr. Dr. Hans Robert Kalbitzer|
|Projects:||Graduiertenkolleg Nichtlinearität und Nichtgleichgewicht|
|Keywords:||Bc, Bacillus caldolyticus; Bs, Bacillus subtilis; CAT, chloramphenicol acetyltransferase; CSP, cold shock protein; dT7, hepta-2'-desoxy-thymidylate; IF, initiation factor; Tm, Thermotoga maritima|
|Subjects:||500 Science > 530 Physics|
|Refereed:||Yes, this version has been refereed|
|Created at the University of Regensburg:||Yes|
|Deposited On:||20 Mar 2007|
|Last Modified:||20 Jul 2011 21:03|