tailieunhanh - Báo cáo khoa học: The influence of cold shock proteins on transcription and translation studied in cell-free model systems

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 sub-tilis, Bacillus caldolyticusand Thermotoga maritima) were studied in an Escherichia colibased cell free expression system on their capability of enhancing protein expression by reduction of mRNA secondary structures. TheE | ềFEBS Journal The influence of cold shock proteins on transcription and translation studied in cell-free model systems Roland Hofweber1 Gudrun Horn1 Thomas Langmann2 Jochen Balbach3 Werner Kremer1 Gerd Schmitz2 and Hans R. Kalbitzer1 1 Institut fur Biophysik und Physikalische Biochemie Universitat Regensburg Germany 2 Institut fur Klinische Chemie und Laboratoriumsmedizin Klinikum der Universitat Regensburg Germany 3 Laboratorium fur Biochemie Universitat Bayreuth Germany Keywords cold shock protein in vitro translation RNA chaperone Correspondence H. R. Kalbitzer Universitat Regensburg Institut fur Biophysik und Physikalische Biochemie UniversitatsstraBe 31 93053 Regensburg Germany Fax 49 941 9432479 Tel 49 941 9432594 E-mail . Received 21 March 2005 revised 7 June 2005 accepted 27 July 2005 doi 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 sub-tilis 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 .