Effect of the 5′ terminal stem–loop structure on the affinity of the 5′-UTR for Hfq and ribosomes

Abstract

<p><b>Copyright information:</b></p><p>Taken from "Both RNase E and RNase III control the stability of mRNA upon translational inhibition by the small regulatory RNA RyhB"</p><p>Nucleic Acids Research 2005;33(5):1678-1689.</p><p>Published online 21 Mar 2005</p><p>PMCID:PMC1069011.</p><p>© The Author 2005. Published by Oxford University Press. All rights reserved</p> () 5′ end-labelled and RNAs were incubated alone (lanes 1 and 6) or with increasing amounts (2-, 4-, 6- and 8-fold molar excess) of Hfq-hexamer (lanes 2–5 and 7–10, respectively), and the resulting mixtures were then analysed on a 6% native gel. The positions of free and RNAs as well as their complexes with Hfq (single and double asterisks, respectively) are indicated. () Differential decrease of 30S ribosome binding to mRNA by and competitor RNA, respectively. The RNA pre-annealed to the 5′ end-labelled primer was incubated with 30S ribosomal subunits in the presence of increasing amounts (1-, 2-, 4- and 8-fold molar excess) of competitor RNAs ( and , respectively). Translation inhibition complex formation was further analysed by primer extension as described in Materials and Methods. () Relative toeprints obtained on RNA [see (B)] using and RNA as competitors, respectively. The relative toeprints (%) were calculated as described by Hartz . () after quantitation of the toeprint and extension signals using the equation: [toeprint signal/(toeprint signal + extension signal)]