PSC2002/L25 Insulin Signalling Flashcards
(46 cards)
Give the 4 protein domains on signalling proteins (tyrosine kinases).
PH - Pleckstrin Homology domain
PTB - phosphotyrosine binding domain
SH2 - Src Homology 2 domain
SH3 - Src Homology 3 domain
Describe the PH domain.
Binds to phosphorylated inositol phospholipid in plasma membrane
Localises the protein to plasma membrane
Describe the PTB domain.
Binds phosphotyrosine (P-Y) residues e.g., in IR
Describe the SH2 domain.
Binds phosphotyrosine residues (P-Y) surrounded by unique protein sequences i.e., SH2 domain in PI3K will not bind to (P-Y) recognised by Grb2
Describe the SH3 domain.
Binds specifically to proline-rich regions i.e., SH3 domain in Grb2 will only bind SOS
Describe the structure of the insulin receptor (IR).
Belongs to Receptor Tyrosine Kinase (RTK) superfamily of 19 families and 55 RTKs
Heterotetramer
190kDa glycoprotein
aaBB subunits held together by disulphide bonds
What are the functions of the a- and B-subunits of the IR?
a-unit: insulin binding domain
B-unit: transmembrane; tyrosine kinase
What is the result of insulin binding to a-units of the IR? (3)
Transphosphorylation of B-units
Activation of tyrosine kinase activity
Phosphorylation of IR generates binding sites for proteins with PTB & SH2 domains
What is the function of the kinase activation loop of the IR?
Activation of tyrosine kinase
Phosphorylation creates binding site for SH2 domain
What is the function of the juxtamembrane domain of the IR?
Phosphorylation creates binding site for PTB domain
What is the function of the C-terminal domain of the IR?
Regulates IR kinase activity and kinase-adaptor protein interactions
What is the function of Ser-Thr in the IR?
Inhibits receptor kinase activity
How many Shc genes are there and what are they comprised of?
3: A, B, C
Shc comprises PTB, CH1, SH2 domains
CH1 (collagen homology domain) has Tyr residues, becomes phosphorylated and binds proteins with SH2 domains e.g., Grb2
Grb2-Sos signals through MAP-Kinase
What is Shc?
SH2-containing adaptor protein
Binds IR through PTB and SH2 domains
How many genes are there for Insulin Receptor Substrate (IRS)?
4
IRS1-4
What is the function of IRS1 and 2 (180kDa)?
Essential for most of insulin’s biological actions in liver, muscle and AT
Describe the structure of IRS.
Contains a PH domain and PTB domain
Enables binding to phosphorylated insulin receptor: IR-Y(P)
Describe the function of Grb2 (growth factor receptor bound protein-2).
Binds to phosphorylated IRS or Shc through SH2
Activates
Uncovers SH3 domain
Binds to SOS (Son of Sevenless) vis SH3 domain
How does SOS act?
As a GDP/GTP exchange factor (GEF)
When is Ras active?
Inactive when bound to GDP
Active when this is exchanged for GTP (by SOS)
Ras-Raf-MEK(MAPKK)-ERK
Drives growth, differentiation, proliferation
Why is IRS1 referred to as a docking protein?
Has 21 potential Y-phosphorylation sites for proteins with SH2 domains
PI3K has 2 SH2 domains
Grb2 has 1 SH2 domain
How many potential S/T-phosphorylation sites does IRS1 have? How are S/T phosphorylated?
30
By components of insulin signalling or by stress kinases that are activated by lipids or inflammation
Describe the 6 steps of the PI-3-Kinase pathway leading to activation of AKT.
- Insulin binds to IR a-subunits to activate IR tyrosine kinase and autophosphorylation of Y
- Recruitment of IRS to IR & phosphorylation of IRS on Ys
- Recruitment of proteins to IRS via SH2 domains & formation of PIP3
- Recruitment of AKT/PKB & PDK1 by PIP3 to plasma membrane
- Dual phosphorylation of AKT
- Phosphorylation of AKT substrates
Describe PI3-Kinase. (4)
Phosphatidylinositol 3-kinase
2 subunit enzyme, p85 (85kDa), p110 (110kDa)
p110 = catalytic subunit
p85 = regulatory unit, contains 2 SH2 domains and 1 SH3
