Growth Factor Receptor Tyrosine Kinases

Question 1

Growth family receptors includes

Answer 1

epidermal growth factors (EGF)
Insulin receptor
Insulin-like growth factor receptor (I)
Platlet driven growth factor (PDGF)

Question 2

Growth family receptor

Answer 2

• mediates cell growth regulation, enzymatic (tyrosine kinase) activity critical for this

Question 3

Ras

Answer 3

small G protein; small polypeptide chain 1/2 size alpha subunit; single AA change keeping it form hydrolyzing GTP back to GDP; had indefinite signal -> cancer

Question 4

3 general features of Receptor Protein Tyrosine Kinases separating them form growth factors

Answer 4

1. An extracellular ligand binding domain (where growth factors initially bind)
2. A cytoplasmic tyrosine kinase domain (these have intrinsic kinase activity)
3. A single membrane spanning domain ( ~20 hydrophobic amino acids) connecting cytoplasmic and extracellular domains

Question 5

Ability of receptor protein tyrosine kinase to phosphorylate proteins

Answer 5

• great regulatory activity
• can regulate protein protein interactions
• can change channel and enzyme activity

Question 6

Tyrosine kinase and cancer

Answer 6

tyrosine kinase activity regulated by C-terminal end; when this is shortened have unregulated activity which -> cancer

Question 7

Oncogenic versions receptor tyrosin-kinases

Answer 7

1. v-ErbB protein truncated EFG receptor in birds; lacks extracellular ligand binding domain and part of C-terminal tail (can’t be down regulated?)
2. Neu/ HER2 oncogene activated by point mutation -> rat neuroblastoma -> breast cancer in humans

Question 8

EGF Receptor

Answer 8

Epidermal growth factor polypeptide hormone which by binding to specific cell surface receptor initiates chain of events culminating in cell growth and proliferation

Question 9

Early and late responses to EGF

Answer 9

early- activation of protein kinase and ion transport activities
late- stimulation of DNA, RNA, and protein synthesis and cell division

Question 10

EGF binding to extracelluarl domain

Answer 10

1. EGF binds to extracellular domain stimulating tyrosine kinase activity of cytoplasmic domain
2. Activated tyrosine kinase phosphorylates itself on C-termina tyrosine residues (phosphorylation is trans)
3. Phosphorylated site= beacon to recruit binding proteins (SH2 domain bind to auto-phosphorylation sites in order to initiate signaling cascades)

Question 11

EGF binding location

Answer 11

far from membrane (50-60 angstroms); different from G coupled protein receptor

Question 12

EGF receptor reatures

Answer 12

• has + charged AAs keep receptor positioned properly, makes sure we receiving and transmitting segments in correct spot
• phosphorylating site tunes down receptor at The
• Lysine necessary for protein binding, have lysin bind ATP
• 7 tyrosine which are sites of autophosphorylation; can recruit diff proteins to bind and start signaling pathways

Question 13

Mechanism of activation of EGF receptor tyrosine kinase activity

Answer 13

EGF-depenent formation receptor dimers, tyrosine kinase domains forms asymmetric dimer one kinase domain serves as the activator, while other kinase domain, upon binding the activator, then becomes activated tyrosine kinase

Question 14

Dimerization and trans-phosphorylation

Answer 14

1. Binding of growth factor to extracellular domain of monomeric EFG receptor exposing dimerization arm
2. Extracellular domain of two receptor monomers come together to form receptor dimer
3. Cytoplasmic domains of 2 receptor monomers interact to form asymmetric dimer
4. trans-phosphorylation where activated tyrosine kinase phosphorylates tyrosine residues within neighboring tyrosine kinase domain (activator being phosphorylated by that which was activated)
5. Recruit cellular proteins to receptor to start signal transduction cascades

Question 15

EGFR degredation

Answer 15

One of auto-phosphorylation sites of EGFR recruits protein c-Cbl which attaches ubiquitin to lysine within cytoplasmic domain of receptor
- ubiquitination of EGFR targets it for degradation in lysosome (ensures EFGR no indefinite signaling)

Question 16

Src homology domains

Answer 16

• common sequence motifs contained by cellular proteins that bind phosphorylated growth factor receptors
• stretches of AA that share sequence ID with regions of src tyrosine kinase

Question 17

SH2

Answer 17

sequences specify for a region for binding to phosphotyrosine-containing proteins (receptors that have been trans-phosphorylated)

Question 18

SH3

Answer 18

• not necessaryy for direct interaction of cellular protein with phosphorylated receptor
• represents distinct region for interacting with stretches of proline residues on other proteins that participate in growth factor-stimulated signaling cascade

Question 19

PI3 kinase

Answer 19

SH2 can plug into regulatory protein of this kinase; this = discovery that you can phosphorylate lipids

Question 20

Ras activation

Answer 20

Grb2 binds specific auto-phosphorylated tyrosine residue on receptor kinase -> 2 SH3 (on Grb2) bind to SoS -> stimulates GDP GTP exchange on Ras(receptors create cascade) -> Ras recruits serine/threonine kinase called Raf to membrane -> kinase cascade -> activation nuclear map kinase called Erk -> Erk translocates to nucleus -> phosphorylation transcriptional regulators and initiation gene expression

Question 21

7 tyr sites of EGF

Answer 21

6 autophosphorylated for signaling 7th binding protein helps with endocytosis b/c leads to ubiquitination

Question 22

verbB

Answer 22

missing 7th binding protein to -> endocytosis therefore -> not shutting off -> cancer

Question 23

Grb2

Answer 23

growth factor binding protein 2; adaptor protein that contains 1 SH2 domain and 2 SH3 domains

Question 24

Neu/ErbB2/HER2 oncogene

Answer 24

all same gene; normal cellular Neu gene product different from transforming Neu from single AA substitution in transmembrane domain; this increases activation of tyrosine kinase

Question 25

Mammary carcinomas

Answer 25

over expression of Neu

Question 26

Heregulin

Answer 26

does not bind HER2/Neu alone; 1st binds either ErbB3 or ErbB4 stimulating heterodimerization of ErbB3 or ErbB4 with HER2/Neu ->heregulin making contact with Neu; cytoplasmic domain ErbB3 serves as activator for tyrosine kinase activity of Neu -> activated Neu trans-phosphorylates ErbB3 -> recruitment of SH2 domain-containing proteins -> initiation of signaling cascades

Question 27

ErbB3

Answer 27

only member of family of receptor tyrosine kinases that lacks kinase activity

Question 28

HER2 regulation

Answer 28

ErbB3 lacks tyrosine kinase activity, serves as receptor for heregulin -> heterodimer formation -> activates neighboring TK -> transphosphorylation

Question 29

Insulin Receptor

Answer 29

starts as dimer with 2 extracellular alpha domains and 2 smaller beta domains; insulin bins alpha subunits and activates B subunit trans-autophosphorylation events (B subunits not properly aligned until insulin binds); Trans-phosphorylation of tyrosine residue in activation loop in active site tyrosine kinase domain stimulates tyrosine kinase activity of each domain -> phosphorylation of protein substrates like IRS-1 protein

Question 30

PDGF

Answer 30

platelet-derived growth factor receptor; dimer brings growth factors together -> signaling
Platlets release PDGF upon activation by thrombin or at sites of blood vessel injury; thought to act upon cells of blood vessel wall; exists as homodimer or heterodimer

Question 31

PDGF structure

Answer 31

extracellular growth factor binding domain and intracellular tyrosine kinase domain separated by single membrane spanning helix
- member of class III receptor tyrosine kinase; possess varibale kinase domain insert sequence (hydrophilic sequence, separates sequences which form ATP binding site and conserved tyrosine autophosphorylation site of kinase domain)

Question 32

kinase domain insert sequence

Answer 32

KI- important for interaction of receptors with with specific intracellular protein targets

Question 33

tyrosine kinase activity of PDGF receptor fully stimulated when

Answer 33

each tyrosine kinase domain phosphorylates a specific tyrosine residue on adjoining tyrosine kinase domain -> additional transphosphorylations of KI regions and binding of phospho-substrates to those phosphorylated sites