Lecture 11: Receptor Tyrosine Kinase
• Receptor tyrosine kinases (RTK)
• Adapter proteins, GRB2
– PTB, SH2, SH3
• Ras and its regulation (GEF, GAP, Sos)
• HERs and human breast cancer
• Ras/MAP kinase pathway operation (Raf, MEK, MAPK), EGF
Receptor tyrosine kinases (RTKs)
• Extracellular (ligand-binding) domain
– Ligands include growth factors and insulin
– Growth can entail cell getting bigger or mitogenic factor that induce a cell to divide.
– Insulin can be considered as a mitogenic factor
• Single transmembrane α-helix (as opposed to GPCR with 7)
• Cytoplasmic domain with intrinsic tyrosine kinase activity which is stimulated by ligand binding
due to receptor dimerization
• Adapter proteins are required
• Ras (downstream effector) acts as a GTPase switch protein to signal further “downstream”
• Aberrant signaling is at root of many human cancers
Activation of RTKs
1. Growth factor binds extracellular domain of 2
2. Each of the transmembrane domains undergo
conformational change that allows dimerization.
3. Dimerization partial activation of the kinase
domain in the cytosolic portion of the RTK.
4. Trans-activation: one kinase domain
phosphorylates and activates the activation lip of
the other RTK.
5. Fully active kinase domain: can self-
phosphorylate the lower tyrosine residues on the RTK.
• Dimerization allows for auto-phosphorylation of the cytoplasmic domain.
– This is important for the recruitment of the Phosphotyrosine.
Phosphotyrosines: serve as docking sites for signal transduction proteins that have SH2 or PTB
domains (adapter proteins)
Page 1 of5 Adapter proteins
Adapter proteins contain unique domains that recognize specific sequences.
Common adapter domains:
• SH2 –src (sarcoma) homology 2 domain
– found in at least 100 human proteins
– binding recognition domain for phosphorylated tyrosines
• PTB – Phosphotyrosine-binding domain.
– Found on multi-docking proteins, serve as docking sites for other signal transduction or
– Binding recognition domain for phosphorylated tyrosines
• IRS-1- Insulin receptor substrate protein
Recruitment of adapter proteins
• Growth factor binding to extracellular domain of the protein à trans-activation: one domain
activated first to activate the other à autophosphorylation moving down on tyrosine residues.
– This is a signal of the proteins that cause recruitment of proteins that recognize
the phosphotyrosines. Those proteins do so through their SH2 and PTB domains.
• When the two proteins with the SH2 and PTB domains get recruited to the RTK kinase domain
and bind to the phosphotyrosines, they may get phosphorylated in some cases.
– Proteins that have PTB domains usually have multi-docking capabilities.
• Multi-docking: IRS-1 when it binds to phosphorylated tyrosine on the RTK, and it is
phosphorylated it acts as a signal for the recruitment of other adapter proteins many of which
will also have SH2 domains.
Discovery of First Oncogene or why is SH2 named like that?
• In 1911 Rous injected a cell free extract from a chicken tumor (sarcoma) into healthy chickens -
all developed tumours
– If you take the sarcoma and grind it all up to kill all the cells and inject it into healthy
chickens, the chickens got tumours.
– This proved that the tumours were not cell-based, it was actually from a virus.
• The transmissible agent was determined to be a retrovirus (Rous Sarcoma Virus)
• In 1980s Bishop-Varmus isolated virus and sequenced it.
– it was found that one of the viral genes encodes a tyrosine kinase (src) protein.
• Sarcomology domains are highly conserved and found in humans (SH2 and SH3)
– The SH2 domain was found in both the viral and human genes.
• Normal version is cellular src (c-Src)
• Viral version is viral src (v-Src) – constitutively active
The Rous Sarcoma virus encoded sarc protein (v-Src) has no regulatory region: It’s missing the
tyrosine residue 527 that is inhibited by phosphorylation.
Page 2 of 5 The kinase encoded is involved in the mutagen activated signaling pathway. So now you don’t need the
mitogen to turn it on the signaling pathway that leads to cell division, v-SRC can just phosphorylate
everything because it is constitutively active.
This is what was causing cancer in the chickens.
If you express v-Src at high levels in a normal human cell, you can make it cancerous.
Ras, a GTPase switch protein
Ras: Monomeric G protein, GTPase superfamily, lipid-anchored protein
• Downstream effector of RTK signaling
• Similar to Gα but smaller, poor GTPase activity(requires GAP)
• Not linked directly to cell-surface receptors
• Ras activity is regulated by GEFs and GAPs.
• Ras protein: 20-30% Human cancers: Ras binds GTP but no hydrolysis, mutation of glycine-
12 which prevents binding GAPs
By itself, Ras has poor GTPase activity. Its regulation is created by GEF (activation) and GAP
RasD is a constitutively active Ras protein, mutant.
• Mutation in cancers where Ras can no longer bind GAP and is constitutively active because it is
permanently hydrolyze to GTP.
EGF-induced Ras activation: step 1
Epidermal Growth Factor (EGF)
1. Ras is inactive (anchored to the membrane by prenylation)
2. EGF binds EGFR as monomers
– EGF binding to each EGFR causes conformational change that leads to dimerization.
3. Trans-activation is prom