Biomedical Engineering Reference
In-Depth Information
b
a
L31
50S
50S
A
P
P
A
E
E
mRNA
L31
30S
30S
+12
-18
Fig. 1.1
Overall view of the 70S ribosome in elongation state. A, P, and E tRNA are shown in
orange
,
red
, and
magenta
, respectively, and 60-mer mRNA (position −18 to +12 visible) is shown
in
gold
. Ribosomal proteins and RNA of the small and large subunits are shown in
light blue
and
violet
, respectively. The new intersubunit bridge formed by the protein L31 is shown in green.
(
a
)
To p
view; (
b
) view from the E site
tRNAs in such a way that the reading frame of the mRNA (each codon consists of
three consecutive nucleotides) is maintained throughout the translation process. The
elongating ribosome contains three binding sites for tRNAs: The aminoacyl (A) site
to which a cognate aminoacyl-tRNA is delivered such that it base pairs with the
appropriate mRNA codon; The peptidyl (P) site where the tRNA carrying the nascent
peptide chain is located. When a cognate aminoacyl-tRNA enters the A site the pep-
tidyl transferase reaction takes place and the peptidyl chain carried by the P tRNA is
added to the aminoacyl tRNA essentially adding one amino acid to the growing
peptide chain; Last is the exit (E) site from which deacylated tRNA that has com-
pleted its role in translation is released.
After each peptide bond reaction, the ribosome must rearrange its contacts with
mRNA and tRNA to allow translocation along the mRNA by a single three nucle-
otides codon. The ribosome controls the positioning of mRNA and tRNAs during
the translation process through a number of direct intermolecular contacts. These
interactions not only help to stabilize the binding of tRNA to the ribosome but are
involved directly in functional processes such as mechanisms for discrimination of
aminoacyl-tRNAs to increase the accuracy of tRNA selection; maintenance of the
correct reading frame to avoid frame-shifting errors; and translocational movement
of the tRNAs and mRNA within the ribosome. We are striving to understand these
mechanisms by studying how the main substrates of protein synthesis such as
mRNA and tRNA interact with the ribosome.
In order to shed light on these issues we recently determined high-resolution
crystal structures of
Thermus thermophilus
70S ribosomal complexes with different
mRNA constructs and naturally modified tRNAs (Fig.
1.1
) (Jenner et al.
2010a,
b
) .
Crystals of the ribosome modeling the elongation state were obtained from ribo-
some complexes prepared with a 60 nucleotides long poly (U) mRNA containing a
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