Co-existing Structures of an mRNA Stability Determinant: The 5′ Region of the Escherichia coli and Serratia marcescens ompA mRNA

Abstract

The structure of untranslated regions of mRNA is thought to play a key role in the degradation of mRNAs by specific RNases. As a model system, in vitro transcripts of the stability determining 5′ non-coding region of bacterial ompA mRNA were investigated by calculation of secondary structure models and by experiments applying the temperature-gradient gel electrophoresis (TGGE). For the theoretical prediction of secondary structures an algorithm was used, which yields the structure of lowest free energy as well as a large set of suboptimal structures. Three structures were predicted to co-exist in similar concentrations under native conditions. They denature in a low temperature transition leading to a unique structure which denatures in a high temperature transition. The prediction of three structures and two transitions could be confirmed experimentally by TGGE. Due to the use of transcripts of different length the conformational transitions could be attributed to distinct parts of the molecules. A pseudoknot structural motif was predicted theoretically, but could not be confirmed experimentally. Comparing ompA transcripts of E. coli and S. marcescens, a conversation of structural features could be shown in spite of a sequence homology of only 63%. Regarding the sequential folding of the transcript after synthesis, a metastable structure is formed first and is converted slowly into structures of lower free energy. The biological implications for in vivo degradation are discussed.