Lecture 11 & 12 (G proteins) Flashcards
Two main classes of signalling G proteins
- Heteromeric
- Monomeric (low molecular weight): Ras superfamily
3 examples of Ras oncogenes
H-Ras, K-Ras, N-Ras
What do Ras genes have in common?
- Encode proteins that are ~188 aa long
- Mw = ~20-25 kDa
- Have GDP and GTP binding activity
Example of a point mutation in cellular Ras gene that changes it to an oncogene
Aspartic acid, valine or cysteine substituted for glycine at amino acid position 12 in Ras
Point mutations in Ras lead to…
the protein being in a permanently active state (GTP-bound)
Oncogenic mutations of Ras either…
(i) decrease the GTPase actvitiy of the Ras protein, or (ii) increase the rate of exchange of bound GDP for GTP
Ras proteins are very frequently activated in __ cancers, particularly __ __
epithelial cancers
pancreatic tumours
~85% of activated Ras in naturally occurring human tumours is __
K-Ras
In recent years, some progress has been made developing drugs that target Ras __
G12C
What does sos do?
It naturally converts the inactive form of Ras to its activated form (sos is a nucleotide exchange protein for Ras)
5 sub-families of the Ras superfamily
- Ras
- Rho
- Rab
- Ran
- ARF (ADP-ribosylation factor)
What is Rho important for?
regulating the actin cytoskeleton
What is the largest sub-family of Ras?
Rab
What is Rab involved in?
the control of vesicular trafficking
What is Ran important for?
movement of proteins in and out of the nucleus
Functional cycle of Ras proteins
Protein active when bound to GTP → protein conformation altered → interacts with effectors → downstream effects → once GTP is hydrolysed and the terminal phosphate is released, the protein returns to its GDP-bound inactivated state
What does Ras-GAP do?
Stimulates the GTPase activity of the G protein to allow Ras to hydrolyse the terminal phosphate from GTP
3 accessory proteins that convert Ras back into its activated state
- GDI (GDP dissociation inhibitor)
- GEF (nucleotide exchange protein)
- REP
Ras proteins are crucial in…
growth factor signal transduction
Domains within Ras proteins that are important for guanine nucleotide binding
G boxes
Which region of G proteins is quite variable
C-terminal region
C-terminal cysteines of the Ras protein are the sites for…
covalent addition of fatty groups like isoprenoids and palmitoyl groups to facilitate membrane association
Where are Ras proteins typically located?
on the plasma membrane
Mammalian Ras has been shown to function upstream of a group of kinases called the…
mitogen activated protein (MAP) kinases
Normally, Ras is activated as a consequence of stimulation of __
RTKs
Do GAP proteins negatively or positively regulate G proteins?
negatively
Example of a GAP that was separately identified as a tumour suppressor gene
NF-1
PTMs of C-terminal CAAX motif that allow monomeric G proteins to tightly associate with membranes
- Farnesylation (isoprenoids)
- Cleavage of -AAX
- Methylation
- Palmitoylation
Best understood growth factor signal transduction pathway
EGF/Ras pathway
Ras directly interacts with __ , and this event is dependent on…
Raf
Ras binding GTP
Grb2 protein complexes with __
sos
Two major pathways downstream of Ras activation
- MAPK
- PI3K
Ral-GEF pathway is activated __ of Ras
downstream
PI3 kinases play an important role as mediators of…
Ras-mediated cell survival and proliferation
PI3K converts __ into __
PIP2 into PIP3
PIP3 binds the __ domain of Akt (PKB)
PH
PI3K can also activate __
Rac (Rho family of Ras superfamily)
2 major effector proteins for Ras
Raf and PI3K
