5 LC Concepts in Molecular Biology 12.10.2022
Translation and protein folding
We have the extensive processing of mRNA in the nucleus, the
depositions of wholemarks like the nuclear cap binding complex
CBC at the 5’ end, the exon junctions complex EJC upstream the
exon-exon junctions, the poly-A binding protein at the poly A tail.
The mRNA has to get to : CBP elF4E and the PABPN1PABPC1
(the poly a binding protein is exchanged to the cytoplasmic part).
mRNA – the closed loop model
Now the cytoplasmic mRNP should be translated. We don’t only have the eIF4E that binds the cap, but
there’s also a second protein eIF4G that binds to the eIF4E.
It was known that the (eIF4) G can interact with the Poly-A binding protein of the cytoplasm, coming the
two ends together making a closed loop model where the 5’ and 3’ can communicate.
it may function as a quality control for incomplete or damaged mRNAs (lacking cap or the polyA tail).
Because, otherwise, how would the 5’ end know what happens to the 3’ end, and vice-versa?
How people see it is that you do have the circular loop model where the rna is
basically circular and the 5’ and 3’ come together, then the ribosome can start
initiating and translating the mrna into the protein. This would somehow
function as a quality control step.
There is some debate, because in the human cells we do have 20.000 mRNA is it
true for all of them?
- Eukaryotic mRNAs are generally monocistronic and
- Initiation usually occurs at the first AUG codon downstream of the 5’
end
- Oreferred start site in mammals is (A/G)XXAUGGKozak sequence
This is important for when you’re designing expression vector and
trasfect an expressed protein in mammalian cells.
It has been observed that if the polyA tail becomes too short the cytoplasm poly
a binding protein cannot be bound anymore; an then of course we cannot have
the interactions with the cap binding factors and this loop will open up and translation is repressed. This is
one of the key findings that short poly a tails are translationally not very active.
If poly a tail becomes too short mRNA is degraded. At least 20% of mRNA are already degraded in the
nucleus, and of course even in the cytoplasm.
mRNA decay in eukaryotes
mRNA in most of the times initiated by deadenylation; so you have this quite long poly-A tail and then
several protein complexes in the cytoplasm that are de-adenylases that specifically remove the poly-A tail
from the 3’ end. If the poly-A tail is to short that it cannot accommodate poly-A binding proteins anymore,
two things can happen:
- can trigger decapping specific complex coming to the 5’ end, removing the cap structure and a
5’-p five prime phosphate exposed. There are 5’->3’ esoribonucleases (Xm1) that will specifically
recognize the 5’-p and degrade in the 3’ direction.
- Exosome + Ski complex in this case exosomes are the micromolecular machines that degrade
rna. mRNA is then degraded from 3’->5’ and even the cap is degraded.
, 5 LC Concepts in Molecular Biology 12.10.2022
TRANSLATION
Ribosomes have to deal with a lot of different mRNA, and
the cytoplasm itself is highly packed full of micromolecular
machines; so the ribosome has to make sure that the
protein folds and that it does not interact with other
factors meanwhile.
Translation is highly regulated and when a protein is
produced, what we called in the nascent chain, it interacts
with various factors along the way until it is totally
produced.
Basically all the time the nascent chain interacts with the ribosome’s exit tunnel; there are some sequences
that very strongly interacts with the exit tunnel and then lead to specifying stoning of the ribosome.
We also have enzymatic modifications.
Early on, as soon as the peptide chain is emerging, enzymes can sit on the ribosome and start to modify the
growing polypeptide; this is something we call targeting factors not all proteins live in the cytoplasm
where they’re made but in other compartments and somehow they need to get there. So the targeting
factors allow proteins to find their destiny.
Chaperons, proteins that help other proteins fold. folding helpers that prevent aggregation of proteins.
Complex assembly as soon when the nascent chain is long enough, that individual domains are already
folding; this can start to interacts with domains or proteins produced in other ribosomes. It has been
shown the already on the translating ribosomes different proteins, also heterocomplexes, come together
and very early start to assembly.
Translation and protein folding
We have the extensive processing of mRNA in the nucleus, the
depositions of wholemarks like the nuclear cap binding complex
CBC at the 5’ end, the exon junctions complex EJC upstream the
exon-exon junctions, the poly-A binding protein at the poly A tail.
The mRNA has to get to : CBP elF4E and the PABPN1PABPC1
(the poly a binding protein is exchanged to the cytoplasmic part).
mRNA – the closed loop model
Now the cytoplasmic mRNP should be translated. We don’t only have the eIF4E that binds the cap, but
there’s also a second protein eIF4G that binds to the eIF4E.
It was known that the (eIF4) G can interact with the Poly-A binding protein of the cytoplasm, coming the
two ends together making a closed loop model where the 5’ and 3’ can communicate.
it may function as a quality control for incomplete or damaged mRNAs (lacking cap or the polyA tail).
Because, otherwise, how would the 5’ end know what happens to the 3’ end, and vice-versa?
How people see it is that you do have the circular loop model where the rna is
basically circular and the 5’ and 3’ come together, then the ribosome can start
initiating and translating the mrna into the protein. This would somehow
function as a quality control step.
There is some debate, because in the human cells we do have 20.000 mRNA is it
true for all of them?
- Eukaryotic mRNAs are generally monocistronic and
- Initiation usually occurs at the first AUG codon downstream of the 5’
end
- Oreferred start site in mammals is (A/G)XXAUGGKozak sequence
This is important for when you’re designing expression vector and
trasfect an expressed protein in mammalian cells.
It has been observed that if the polyA tail becomes too short the cytoplasm poly
a binding protein cannot be bound anymore; an then of course we cannot have
the interactions with the cap binding factors and this loop will open up and translation is repressed. This is
one of the key findings that short poly a tails are translationally not very active.
If poly a tail becomes too short mRNA is degraded. At least 20% of mRNA are already degraded in the
nucleus, and of course even in the cytoplasm.
mRNA decay in eukaryotes
mRNA in most of the times initiated by deadenylation; so you have this quite long poly-A tail and then
several protein complexes in the cytoplasm that are de-adenylases that specifically remove the poly-A tail
from the 3’ end. If the poly-A tail is to short that it cannot accommodate poly-A binding proteins anymore,
two things can happen:
- can trigger decapping specific complex coming to the 5’ end, removing the cap structure and a
5’-p five prime phosphate exposed. There are 5’->3’ esoribonucleases (Xm1) that will specifically
recognize the 5’-p and degrade in the 3’ direction.
- Exosome + Ski complex in this case exosomes are the micromolecular machines that degrade
rna. mRNA is then degraded from 3’->5’ and even the cap is degraded.
, 5 LC Concepts in Molecular Biology 12.10.2022
TRANSLATION
Ribosomes have to deal with a lot of different mRNA, and
the cytoplasm itself is highly packed full of micromolecular
machines; so the ribosome has to make sure that the
protein folds and that it does not interact with other
factors meanwhile.
Translation is highly regulated and when a protein is
produced, what we called in the nascent chain, it interacts
with various factors along the way until it is totally
produced.
Basically all the time the nascent chain interacts with the ribosome’s exit tunnel; there are some sequences
that very strongly interacts with the exit tunnel and then lead to specifying stoning of the ribosome.
We also have enzymatic modifications.
Early on, as soon as the peptide chain is emerging, enzymes can sit on the ribosome and start to modify the
growing polypeptide; this is something we call targeting factors not all proteins live in the cytoplasm
where they’re made but in other compartments and somehow they need to get there. So the targeting
factors allow proteins to find their destiny.
Chaperons, proteins that help other proteins fold. folding helpers that prevent aggregation of proteins.
Complex assembly as soon when the nascent chain is long enough, that individual domains are already
folding; this can start to interacts with domains or proteins produced in other ribosomes. It has been
shown the already on the translating ribosomes different proteins, also heterocomplexes, come together
and very early start to assembly.
