Investigating RNAs Involved in Translational Control by NMR and SAXS - Biophysical Approaches to Translational Control of Gene Expression - page 159

Fig. 8.13 Solution structure of the TCV RBSE (Zuo et al. 2010 ) . ( a ) Secondary structure of the
RBSE. Long-range base-pairing interactions are indicated by dashed lines . ( b ) The solution struc-
ture of the TCV RBSE (PDB ID 1KRL) as determined by a combination of SAXS and NMR
reveals a twisted T-shape. ( c ) Despite the lack of similarity in secondary structure, the TCV RBSE
( purple ) resembles a tRNA ( gray ) in its three-dimensional fold

(Fig. 8.13a ). The RBSE assists in recruitment of the large ribosomal subunit and
contains a large asymmetric loop that is essential for the coordination of translation
of the viral proteins with genome replication. This study utilized the G2G software
package that first models the individual A-form helices in the RNA, then aligns the
helices by fitting them to the RDC data (Wang et al. 2009 ) . Linker sequences are
then modeled into the aligned helices to create a complete model. The resulting
model was iteratively refined against both SAXS and NMR data in XPLOR-NIH
(Fig. 8.13b ). Interestingly, the RBSE RNA mimics a tRNA molecule in overall
shape (Fig. 8.13c ), despite the fact that the RBSE secondary structure is quite dis-
tinct from the typical cloverleaf tRNA secondary structure (Fig. 8.13a ) (Zuo et al.
2010 ). Thus, the TCV RBSE likely binds to the ribosome via its ability to mimic a
cellular tRNA.

8.2.2.3 RNA Thermometers

RNA thermometers regulate translation initiation through temperature-dependent
RNA structure formation. Rinnenthal et al . have investigated a stem-loop structure
found in the 5¢ UTR of the Salmonella fourU RNA that base-pairs to the Shine-
Dalgarno sequence at low temperatures, preventing ribosome recruitment
(Rinnenthal et al. 2010 ). At high temperatures, the structure unfolds, exposing the
Shine-Dalgarno sequence and allowing translation to occur. To investigate the
temperature-dependent dynamics of this structure, NMR was used to measure the ther-
modynamic stability of individual base-pairs (Rinnenthal et al. 2010 ) . Thermodynamic
stability of the base-pairs was quantified by measuring the imino 1 H exchange rates