Colocation and Autoinhibition Flashcards
Why is colocation important?
The function of a protein is not only defined by its activity but also by its location (eg isoforms of RAS)
What are the three isoforms of RAS?
K-RAS and N/H-RAS
What is the difference between the three isoforms of RAS?
K-RAS has a farnesyl chain and is found only at the membrane.
N/H-RAS have a palimoytyl chain and are found both at the Golgi and at the cell membrane
What are PH domains?
PH domains bind to the membrane and inositol phsophates. Attach components to the membrane in a defined orientation
What is an example of a protein we have studied that utilises a PH domain for colocalization?
Protein kinase B - recruited and activated via PIP3 PH domain
What molecules would you find to be increased in a lipid raft?
cholesterol, sphingolipids and saturated fatty acids.
(also prenylated proteins, dually acetylated proteins and GPI anchored proteins but plz do not mark me down if I don’t say these just an extra juicey juice if I need it)
When is autoinhibition of a molecule most likely to occur?
When there are domains connected by intramolecular binding
What is the basic mechanism of autoinhibition in EGF receptor?
Activation lip in this receptor has conformation that blocks the active site when in an unphosphorylated state.
How is the kinase domain separated from the transmembrane domain in EGFR?
Via the juxtamembrane segment
in the monomeric state, how is the conformation of EGFR affected by the juxtamembrane segment?
1 part of the juxtamembrane segment binds the upper or N-LOBE of the kinase domain.
This causes a conformational change which means the activation lip sits in the active site, blocking its activity
How does dimerization remove the autoinhibition of EGFR?
Dimerisation causes and assymetric dimer to form, so that one kinase (the activator) binds the juxtamembrane segment of the other kinase (the receiver)
This causes a conformational change in the N lobe of the receiver, pushing the activation lip out of the active site.
Can now function as a kinase
How is SOS autoinhibited?
SOS is recruited to the membrane by Grb2.
Once it is at the membrane it binds a different RAS-GDP to the final one it will activate.
This binding activates SOS as a GEF.
The newly activated RAS-GTP will bind SOS and SOS becomes even better GEF.
PH domain in SOS will then bind PIP2 in membrane increasing activity of SOS even further.
How is SRC kinase autoinhibited?
modular structure allows autinhibition
SH2 domain binds P-tyr near c-terminus
SH2-P-Tyr fixes the linker which contains a proline rich sequence
SH3 clamps linker shut and active site is blocked
What are the three ways autoinhibited SRC kinases can be activated?
De-phosphorylating the P-tyr
Binding of SH2 to ‘better’ P-tyr, eg substrate
Binding of SH3 to better polyproline
How is the autoinhibition of AB1 kinase different to SRC kinase?
SH2 domain binds to the back of the kinase not to a phosphorylated tyrosine residue.
Structure is locked by N terminus peptide
