BIO120H1 Lecture Notes - Lecture 6: Contortion, Motor Protein, Hydrophobe
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Thursday, January 22, 2008
- Today we continue with sorting of proteins to the endoplasmic
reticulum, another form of transmembrane transport.
- Recall there are 2 types of ER: the rough ER (actively carrying out
protein synthesis with ribosomes associated with it – the ribosomes are
the things that give it the rough appearance) & smooth ER. We are going
to focus on the rough ER today.
- Proteins are sorted to the ER through transmembrane transport – recall
this is the 2nd type of transport that we covered in the last lecture – the 1st
type is gated-transport & then we covered transmembrane transport. Once
proteins enter the ER from the cytosol & from there, they can be further
sorted to other compartments such as the Golgi, lysosomes or even
secreted to the cell exterior by vesicular transport.
- Today we’re going to look at how proteins get from the cytosol into the
ER by transmembrane transport.
- ER is not a static structure, it is actually changing all the time in terms
of its location in the cell & its composition in the cell. This video nicely
- Video: This would be an ER network here & just take a look at how that
network changes just as you’re looking at it over time.
- Description: The ER is a highly dynamic network of interconnected
tubules that span the cytosol of the eucaryotic cell, like a spider’s web.
The network is continually reorganizing with some connections being
broken while new ones are being formed. Motor proteins moving along
microtubules can pull out sections of ER membranes to form extended
tubules that then fuse to form the network.
- This is just to demonstrate how dynamic this whole process – ER is
very dynamic & proteins are coming into it at multiple locations where
there is the rough ER with ribosomes associated with it.
Synthesis & modification of proteins (covered in this lecture)
Synthesis of lipids (covered in a later lecture)
Have ER signal sequence
- The proteins that are sorted to the ER all have a signal sequence which
is present on the protein so this would be the mRNA here (in the
diagram), a ribosome associates with it & starts to translate that protein &
the ER signal sequence comes out at the N-terminus – the 1st part of the
protein that comes out of the ribosome so it’s an N-terminal endoplasmic
reticulum signal sequence – ER signal sequence.
- The types of proteins that have an ER signal sequence are both soluble
proteins & transmembrane proteins & these are many proteins destined
for the Golgi, secretion outside the cell & also to the lysosomes. Also
some proteins are destined to stay in the ER.
Hydrophobic AAs at N-terminus
- mRNA comes out of the nucleus & then ribosomes associate with the
mRNA & to translate that mRNA into protein. Normally, what occurs is
that multiple ribosomes will bind to an mRNA molecule & you’ll get
polyribosomes in the cytosol associated with that mRNA & each one of
these ribosomes can be translated into protein from this one mRNA
molecule. That’s where all proteins start, most of them, in the cytosol.
Some of them have an ER signal sequence that emerges at the N-
terminus. This sequence that emerges is a hydrophobic AA sequence
without any characteristic specific AA sequence, it’s mainly this
hydrophobicity that gives it its characteristic ER signal sequence. For an
ER protein, as it’s being translated from the ribosome, the ER signal
sequence will emerge 1st at the N-terminus.
- Once the ER signal sequence emerges from the ribosome, this whole
complex is then directed to the ER membrane. As soon as the ER signal
sequence emerges, it’s directed to the ER membrane, then the protein that
is destined to go into the ER is directed through transmembrane transport
in a co-translational process into the ER so the ribosome continues to
translate the protein as it’s being funneled into the ER.
- How this occurs is that there is actually this signal recognition particle
cycle that occurs, that recognizes this ER signal sequence as it’s
emerging from the ribosome so there’s a signal recognition particle that
will bind to the ER signal sequence that will pause translation & then that
whole complex with the signal recognition particle will be directed to a
signal recognition particle receptor that’s on the ER membrane then
translation will resume & the protein will be integrated into the ER in a
co-translational process as the protein is being translated off the
- There are 2 major components to direct the emerging peptide with the
signal sequence to the ER: the signal recognition particle also referred to
SRP & the single recognition receptor SRP receptor.
- Diagram: Here is the ribosome, the mRNA & the protein that’s destined
for ER is being translated – the first thing to emerge is this N-terminal ER
signal sequence – this will be bound by the SRP shown in the diagram &
2nd this SRP will bind to the SRP receptor that is found in rough ER
membranes – that will bring this entire complex to the ER. Both of these
proteins have GTPase domains & they’re normally bound to GTP in this
phase of the process where they’re bringing the ribosome & the newly
synthesized protein to the ER. So they are both bound to the GTP so both
the SRP & the SRP receptor have GTPase domains & are bound to GTP
at this state.
- Normally the SRP & the ribosome have a very low affinity for each
other. When the ribosome is free without a protein coming out of it with
the signal sequence, there is very low affinity b/w the 2 but when the SRP
encounters a ribosome that has a protein with a ER signal sequence
coming out of it then that becomes a very high affinity interaction & the
SRP will bind to the signal sequence & then it can guide the protein &
ribosome to the ER by binding to this SRP receptor.
Prevent diffusion of ions, small molecules
- In the ER, there is this protein translocator through which the newly
synthesized protein will be funneled through. So you can think of it kind
of like a tube of tooth-paste where the protein coming out would the
toothpaste coming out of this ribosome which would kind of be the tube
of toothpaste squeezing the protein out & injecting it into this protein
translocator into the ER.
- Diagram: The ribosome is brought to the protein translocator once it’s
bound to the SRP receptor. Here is the ribosome with the newly
synthesized peptide, the ER sequence here has bound to the SRP, SRP is
bound to SRP receptor & now this entire complex is brought to this
- The translocator channel is also called the translocon & it’s a gated
channel meaning that it’s not always open – it’s kind of shown in the
diagram as a plug but this will actually open once the ribosome & the ER
signal peptide binds to the translocator. So what happens then is these 2,
the SRP & the SRP receptor, bring the ribosome & the newly synthesized
peptide to the translocator & the binding of the ribosome & the ER signal
peptide to the translocon opens it up & allows the protein to be
synthesized through the translocon. Now the ribosome forms a very tight
seal with the translocator & this is important so it prevents the diffusion
of ions & small molecules out of the ER.
- Once this happens, how does this dissociation occur? What’s known is
the SRP & the SRP receptor hydrolyze their GTP to GDP & inorganic
phosphate & this somehow releases the complex so the SRP will be
released into the cytoplasm & the SRP receptor will be released back into
the ER membrane. It’s not known exactly what triggers this GTP
hydrolysis at this time but what’s known is that GTP hydrolysis precedes
the dissociation of the entire complex.
Cleaves ER signal sequence
Translocator gated in the 2nd direction
- Examples of how proteins are inserted into the ER.
- Example of a soluble protein – this has no transmembrane domains &
the 1st thing is that these proteins have an ER signal sequence at the N-
terminus (again). All the translation is occurring in the cytosol & what is
not shown here is the SRP & the SRP receptor but what you have to
assume what has just happened is that the SRP & the SRP receptor have
brought this newly synthesized protein to the translocon.