Format

Send to

Choose Destination
Mol Cell. 2014 Jul 3;55(1):148-55. doi: 10.1016/j.molcel.2014.05.017. Epub 2014 Jun 19.

Direct evaluation of tRNA aminoacylation status by the T-box riboswitch using tRNA-mRNA stacking and steric readout.

Author information

1
National Heart, Lung and Blood Institute, 50 South Drive, MSC 8012, Bethesda, MD 20892-8012, USA.
2
National Heart, Lung and Blood Institute, 50 South Drive, MSC 8012, Bethesda, MD 20892-8012, USA. Electronic address: adrian.ferre@nih.gov.

Abstract

T-boxes are gene-regulatory mRNA elements with which Gram-positive bacteria sense amino acid availability. T-boxes have two functional domains. Stem I recognizes the overall shape and anticodon of tRNA, while a 3' domain evaluates its aminoacylation status, overcoming an otherwise stable transcriptional terminator if the bound tRNA is uncharged. Although T-boxes are believed to evaluate tRNA charge status without using any proteins, this has not been demonstrated experimentally because of the instability of aminoacyl-tRNA. Using a simple method to prepare homogeneous aminoacyl-tRNA, we show that the Bacillus subtilis glyQS T-box functions independently of any tRNA-binding protein. Comparison of aminoacyl-tRNA analogs demonstrates that the T-box detects the molecular volume of tRNA 3'-substituents. Calorimetry and fluorescence lifetime analysis of labeled RNAs shows that the tRNA acceptor end coaxially stacks on a helix in the T-box 3' domain. This intimate intermolecular association, selective for uncharged tRNA, stabilizes the antiterminator conformation of the T-box.

PMID:
24954903
PMCID:
PMC4104367
DOI:
10.1016/j.molcel.2014.05.017
[Indexed for MEDLINE]
Free PMC Article

Supplemental Content

Full text links

Icon for Elsevier Science Icon for PubMed Central
Loading ...
Support Center