Nucleic Acids Research Advance Access originally published online on June 18, 2007
Nucleic Acids Research 2007 35(13):4331-4346; doi:10.1093/nar/gkm439
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Nucleic Acids Research, 2007, Vol. 35, No. 13 4331-4346
© 2007 The Author(s)
This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/2.0/uk/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.
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In vitro analysis of the interaction between the small RNA SR1 and its primary target ahrC mRNA
1AG Bakteriengenetik, Friedrich-Schiller-Universität Jena, Philosophenweg 12, Jena D-07743, Germany and 2Max F. Perutz Laboratories, Department of Microbiology and Immunobiology, University Departments at the Vienna Biocenter, Dr Bohrgasse 9/4, 1030 Vienna, Austria
*To whom correspondence should be addressed. Tel: +49 3641 949570/571; Email: Sabine.Brantl{at}rz.uni-jena.de
Received April 20, 2007. Revised May 16, 2007. Accepted May 16, 2007.
Small regulatory RNAs (sRNAs) from bacterial chromosomes became the focus of research over the past five years. However, relatively little is known in terms of structural requirements, kinetics of interaction with their targets and degradation in contrast to well-studied plasmid-encoded antisense RNAs. Here, we present a detailed in vitro analysis of SR1, a sRNA of Bacillus subtilis that is involved in regulation of arginine catabolism by basepairing with its target, ahrC mRNA. The secondary structures of SR1 species of different lengths and of the SR1/ahrC RNA complex were determined and functional segments required for complex formation narrowed down. The initial contact between SR1 and its target was shown to involve the 5' part of the SR1 terminator stem and a region 100 bp downstream from the ahrC transcriptional start site. Toeprinting studies and secondary structure probing of the ahrC/SR1 complex indicated that SR1 inhibits translation initiation by inducing structural changes downstream from the ahrC RBS. Furthermore, it was demonstrated that Hfq, which binds both SR1 and ahrC RNA was not required to promote ahrC/SR1 complex formation but to enable the translation of ahrC mRNA. The intracellular concentrations of SR1 were calculated under different growth conditions.
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