KEY TERMS:
- The context of a codon in mRNA refers to the fact that neighboring sequences may change the efficiency with which a codon is recognized by its aminoacyl-tRNA or is used to terminate protein synthesis.
- IF-2 binds the initiator fMet-tRNAf and allows it to enter the partial P site on the 30S subunit.
The meaning of the AUG and GUG codons depends on their context. When the AUG codon is used for initiation, it is
read as formyl-methionine; when used within the coding region, it represents
methionine. The meaning of the GUG codon is even more dependent on its location.
When present as the first codon, it is read via the initiation reaction as
formyl-methionine. Yet when present within a gene, it is read by Val-tRNA, one
of the regular members of the tRNA set, to provide valine as required by the
genetic code.
How is the context of AUG and GUG codons interpreted? Figure 6.14 illustrates the decisive role of the ribosome,
in conjunction with accessory factors.
In an initiation complex, the small subunit alone is bound
to mRNA. The initiation codon lies within the part of the P site carried by the
small subunit. The only aminoacyl-tRNA that can become part of the initiation
complex is the initiator, which has the unique property of being able to enter
directly into the partial P site to recognize its codon.
When the large subunit joins the complex, the partial
tRNA-binding sites are converted into the intact P and A sites. The initiator
fMet-tRNAf occupies the P site. and the A site is available for entry
of the aminoacyl-tRNA complementary to the second codon of the gene. The first
peptide bond forms between the initiator and the next aminoacyl-tRNA.
Initiation prevails when an AUG (or GUG) codon lies within a
ribosome-binding site, because only the initiator tRNA can enter the partial P
site generated when the 30S subunit binds de novo to the mRNA. Internal
reading prevails subsequently, when the codons are encountered by a ribosome
that is continuing to translate an mRNA, because only the regular
aminoacyl-tRNAs can enter the (complete) A site.
Accessory factors are critical in controlling the usage of
aminoacyl-tRNAs. All aminoacyl-tRNAs associate with the ribosome by binding to
an accessory factor. The factor used in initiation is IF-2 (see 6.4 Initiation in bacteria needs 30S
subunits and accessory factors), and the corresponding factor used at
elongation is EF-Tu (see 6.10 Elongation factor Tu loads aminoacyl-tRNA into the A site).
The initiation factor IF-2 places the initiator tRNA into
the P site. By forming a complex specifically with fMet-tRNAf, IF-2
ensures that only the initiator tRNA, and none of the regular aminoacyl-tRNAs,
participates in the initiation reaction. Conversely, EF-Tu, which places
aminoacyl-tRNAs in the A site cannot bind fMet-tRNAf , which is
therefore excluded from use during elongation.
An additional check on accuracy is made by IF-3, which
stabilizes binding of the initiator tRNA by recognizing correct base pairing
with the second and third bases of the AUG initiation codon.
Figure 6.15 details the series of events by which IF-2
places the fMet-tRNAf initiator in the P site. IF-2, bound to GTP,
associates with the P site of the 30S subunit. At this point, the 30S subunit
carries all the initiation factors. fMet-tRNAf. then binds to the
IF-2 on the 30S subunit. IF-2 then transfers the tRNA into the partial P site.