Eukaryotic translation elongation closely parallels prokaryotic mechanisms, primarily involving two key elongation factors (eEF1A and eEF2) that facilitate tRNA binding and ribosome translocation, with specific eukaryotic nuances like Hoogsteen base pairing and a dipthamide modification on eEF2.
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Typically we think of ukareotic
translation elongation to be very
similar to the one that we have seen in
procariats. So you should definitely
watch that video before continuing this
one. The link to the complete
translation playlist is down in the
description as well as in the cards on
the top right corner. In proaryotic
translation elongation, we saw two main
elongation factors EFTU and EFG that
were involved in the moving of the
ribosome on the mRNA. In ukareiats,
there are two elongation factors that
are equivalent to the ones found in
procaryots. So EFTU has an orthologue
named ukareotic elongation factor 1A and
the EFG has an orthologue named
ukareotic elongation factor 2. In this
video we will discuss these two
elongation factors and how they work and
we will also see how hoostine base
pairing is involved in the translation
process. So we start where we left off
at the end of the initiation which is
the ads ribosome complex where the aug
start codon is bound by the initiator
tRNA that carries the methionine. So you
have the P site occupied by the transfer
RNA and the A site and the E site are
both empty. And as we saw in proariats,
the 60S subunit contains the factor
binding region. And this complex moves
on to the next step where the amino
acillated tRNA has to position itself at
the respective codon at the A site. And
this amino acillated tRNA is carried by
the elongation factor 1A which means
that the transfer RNA and the elongation
factor form a turnary complex. This
turnary complex therefore has the
elongation factor 1A which just like
EFTU carries a GTP and you also have the
transfer RNA in this complex. Let's
actually draw out this terinary complex.
The elongation factor 1A binds near the
threep prime end of the transfer RNA
close to the amino acid. Before we move
on, I just wanted to highlight that this
elongation factor 1A. The promoter of
this gene that encodes this factor is
quite frequently used in cloning where
some ectopic gene which has to be
expressed in mamalian cells is actually
driven by the promoter of EF1A.
And this factor EF1A is also sometimes
called elongation factor 1 alpha or
simply E F1 alpha. Now let's draw out
the complex that results when elongation
factor alpha with its transfer RNA binds
the codon at the A site. The elongation
factor 1A just like EFTU in procaryots
is positioned near the factor binding
region. And just for your note, the E
site is still not occupied by anything.
One detail worth mentioning at this step
is that the codon and anti-codon pairing
is partially controlled by the 18S
ribosomal RNA which is part of the 40S
ribosome subunit. Specifically the ATS
ribosomal RNA stabilizes the codon and
anti-codon pairing and it is the
secondary structure that is formed by
the ribosomal RNA and specifically at
the adnines in that secondary structure
that reach out into the major and minor
grooves of the anti-codon codon pairing
to help in the overall stabilization. So
if we simply draw this out, what we
notice is that when the pairing of mRNA
and tRNA is established, the 18S
ribosomal RNA is also trying to somehow
pair within this region at the A site.
It is not complimentary pairing, but it
is an interaction of adinine bases into
the codon and anti-codon pairs. And this
looks a lot like the triple helix
hydrogen bonding, which is based on the
principles of non-cononical base
pairing. one of which is known as the
Hoostein base pairing. These base
pairing mechanisms are special and
probably deserve to be addressed in a
separate video, but you should be
mindful of such subtleties in biological
processes. So when the anti-codon and
codon pairing is established and
stabilized, the SRL region in the factor
binding region activates the GTPAS and a
phosphate is released from the EF1A
factor. This causes the elongation
factor 1A to be in a complex with GDP
and as a result the EF1A is released. In
the case of EF1A, the conversion of GTP
to GDP actually causes a structural
rotation in the elongation factor 1A
complex. So it is the rotation which
releases the elongation factor 1A out of
this complex and not simply because it
just loses the affinity. And we know
that this is the rotation because some
specific translation inhibitors are
known to target the region which causes
the rotation of the elongation factor
1A. Dademian B is one such known
inhibitor. Anyways, the proper pairing
of the codon and anti-codon, the
accommodation mechanism that we talked
about in procaryots to position the tRNA
and the release of the elongation factor
1A brings the amino acid on a site tRNA
close to the amino acid on the P site.
And now we can draw out the next step.
And you notice that the amino acid on
the tRNA's at the pite and a site are
very close to each other. The e site is
still empty. All right. Now we also know
from the translation introduction video
that the 60S subunit contains a 28S
ribosomal RNA and like the 23S ribosomal
RNA and procariats the 28S ribosomal RNA
is responsible for peptide bond
formation in the ukareots. Now sometimes
under special condition the peptide bond
formation may require additional
translation elongation factors which in
ukarots is known as elongation factor 5a
which binds at the e site but largely
its main role appears to be in ensuring
efficiency and fidelity of the ribosome.
So mostly it has regulatory functions.
Oftentimes proline repeats need help
from elongation factor 5A but otherwise
it is not as essential in procariats.
Similar function is done by elongation
factor P. The overall process and the
mechanism of peptide bond formation is
similar in both procariots and ukareots
which we talked about in the proarotic
elongation process and this was
described by the proton shuttle and the
proton wire mechanism. If you're
interested in these mechanism, you
should definitely check out the
translation elongation step in
procariats for details. The link to that
video is down in the description.
Following the peptide bond formation, we
can draw out the resulting structure.
And what we notice, unsurprisingly, is
that the methunine from the pite tRNA is
transferred onto the tRNA at the A site.
The E site is still empty. Essentially
all we have done is formed a peptide
bond between the two amino acids. And
now this complex is ready for the next
step which is called transllocation. And
this step of transllocation needs the
second elongation factor elongation
factor 2 which just like EFG in
proariots comes with a GTP attached to
it. But there's a small difference in
ukareotic orthologue because EF2
contains a post translational
modification at one end of the protein
which is known as diptomide which is
just a chemical modification of the
histadine amino acid on the protein and
this dipomide modification is often
times required for breaking the 18S
ribosomal RNA interaction with the mRNA
and the tRNA at the A site. So in this
ATS ribosome the EF2 positions itself at
the factor binding region where the
daptoide modification is close to the
codon anti-codon pairing region and as
we saw in the procariats the aight tRNA
because it has the amino acid chains
wants to naturally move to the pite
because there's a protein exit channel
which is very close to the pite. So this
causes the tRNA to move and as a
feedback the 40S subunit rotates and
this causes the chain to move into the
protein exit channel. The energy for
this movement is provided by the
phosphate bond present in the GTP of the
elongation factor 2. So after
transllocation, the AUG start codon
which was at the P site now moves into
the E site and the A site codon moves
into the P site and as a result the
associated transfer RNA also move with
them because they're paired with the
codons and now the amino acid chain can
go into the protein exit channel. So the
ribosome has moved by one codon and the
AITE is occupied by the EF2 GDP complex.
This transllocated ribosome now moves to
its original position and the 4DS
subunit rotates back which causes the
EF2 GDP which anyways has low affinity
because of its loss of phosphate from
the GTP to be released and the eite tRNA
is also released. So in the next step
the P site is the only occupied site
where the amino acid chain is linked to
the transfer RNA and both E and A sites
are empty. This situation is exactly
like the starting ADS complex except
that the amino acid chain is a bit
longer and this is ready for the next
amino acillated transfer RNA to be
brought in by the elongation
[clears throat] factor 1A. And with each
cycle of translation elongation, the
amino acid chain will continue to grow
until the A site encounters a stop
codon. And when a stop codon is
encountered, the ribosome under goes the
termination step which will be the next
video. Now before you leave, here's a
cool interesting twist in ukarotic
translation. In fungi specifically, they
have an elongation factor known as
elongation factor 3 that binds the eite
tRNA and causes it to release. This is
special to only fungi. Us humans and
other higher order ukareots don't have
this factor. It's interesting how many
things in translation are conserved
across species but fungi have their own
little trick. And this wraps up our
discussion for translation elongation
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