L18 - Receptor Tyrosine Kinases

Question 1

How many families of RTKs

Answer 1

16 families

Question 2

Ligands can be of what two broad calss

Answer 2

Hi or lo affinity

Question 3

What is the structure of the EC domain of the RTK

Answer 3

These vary greatly

Question 4

What activity to the IC domain of the RTK have

Answer 4

Kinase activity

Question 5

How many TMD of the RTK

What is the strcutre, what is the question this produces

Answer 5

1

Lacks structure

Question: How is the ligand binding transduced to the inside of the cell

Question 6

RTK ligand has the abiliy to …

What can this do

Answer 6

DImerise and facillitates receptor dimerisation

May also reorient existing receptor oligomers

Question 7

Describe what happens to the RTK once the ligand is bound

Answer 7

Kinase domains phosphorylate each other

Question 8

Desribe the effects (3) once the kinase domains have phosphorylated each other

Answer 8

Increases the activity of the kinase
Stabilises the receptor in the active state
Casues the kinase to phosphorylate other tyrosines in the receptor to create a docking site

Question 9

Tools for RTK analysis exploit …

Answer 9

Dimerisation

Question 10

Describe how a dominatn geative RTK would be produced

Answer 10

Using genetic engineering to make DNA encoding a receptor which is MUTATED IN THE KINASE DOMAIN

Expressed at high levels and poisons the endogenous receptor

Question 11

How is a constiuently active RTK produced

Answer 11

Use genetic engineering to make an RTK lacking the homodimerisation domain

DNA expressed at normal levels and is ligand independent

Question 12

What are HSPGS

Answer 12

Multifunctional extracellular matrix components

Question 13

Describe the types of protein core in HSPGs (3)

Answer 13

Transmembrane
Tethered
Secreted

Question 14

What happens to the sugars on HSPGs

Answer 14

They can be moified

Question 15

What is one of the most common modifcations on the HSPG sugar

Answer 15

Sulphation

Question 16

What does modificaiton of the HSPG sugar result in

Answer 16

Results in a code that creates binding sites for specific proteins

Question 17

What is a protein which commonly binds to HSPG

Answer 17

FGF2

Question 18

Example of a secreted HSPG

Answer 18

Perlecan

Question 19

Example of a tethered HSPG

Answer 19

Glypican

Question 20

Example of a transmembrane HSPG

Answer 20

Syndecan

Question 21

What occurs when FGF binds to its receptors

Answer 21

FGF and its receptor form a complex with HSPGs

FGF first forms oligomers of HSPGs

Question 22

How does the activated receptor transduce the signal

Answer 22

Activated receptor recruits other proteins to the membrane

These protiens bind to phosphotyrosines

Question 23

RTK have two binding sites - name them

Answer 23

Binding site for phosphotyrosine

Binding site for amino acid side chain

Question 24

The SH2 domain in the RTK recongises which sequence

Answer 24

Phosphtyrosine - glutamic acid - glutamic acid - isoleucine

Question 25

What 2 branches are there in the RTK pathway

Answer 25

RAS | PI3 kinase

Question 26

What 3 proteins can bind to the PDGF receptor

Answer 26

PI3 kinase GTPase activating protein (GAP) Phospholipase C-gamma

Question 27

PI3 kinase and PLC-gamma are part of which pathway

Answer 27

Inositol lipid pathway

Question 28

What is the GTpase activating protein part of which pathway

Answer 28

RAS/MAP kinase pathway

Question 29

What are some of the family members of the Ras family

Answer 29

H-RAS K-RAS N-RAS Rheb Rep1

Question 30

Functions of H-Ras K-Ras and N-Ras

Answer 30

Relays signals from RTKs

Question 31

Functions of Rheb

Answer 31

Activates mTOR to stmulate cell growth

Question 32

function of Rep1

Answer 32

Activated by a cAMP dependent GEF | Influences cell adhesion by activating integrins

Question 33

Family members of Rho

Answer 33

Rho Rac cdc42

Question 34

functions of rho rac and cdc42

Answer 34

Relay signals from surface receptors to the cyotskeleton and elsewhere

Question 35

What binds causing the coupling of the receptor to inactive ras

Answer 35

GRB2 and Sos(GEF)

Question 36

What is the effect of GRB2 and Sos(GEF) bidning

Answer 36

Couples the receptor to inactive ras

Question 37

What does Sos promote What type of molecule is it

Answer 37

Promotes dissociation of GDP from Ras GEF (gunaine nucelotide exhcnage factor)

Question 38

What occurs after GDP has dissociated

Answer 38

GTP binds to Ras | Sos dissociates from the now active Ras

Question 39

Describe the MAP kinase pathway which is downstream of the activated Ras

Answer 39

GTP-Ras activates MAP-KKK MAP-KKK --> MAP-KK (phosohrylation consuming ATP) MAP-KK --> MAP-K (phosohrylation consuming ATP) MAP-K can then phosphorylate target proteins

Question 40

What are the two targets for MAP-K what does phosphorylation at each cause

Answer 40

protein - changes protein activity Gene regulatory proteins - changes in gene expression

Question 41

MAP-K pathways is an example of a __________ | What are the properites of this

Answer 41

Cascade Rapid and transient response

Question 42

What occurs at each stage of the MAP pathway

Answer 42

Amplificaiton

Question 43

What methods may be used to study signalling

Answer 43

Visualisation or detection of interations Chemical inhibitors Misexpression or overexpression Genetic methods

Question 44

what types of visualisation methods may be used to study signalling

Answer 44

``` Biochemical methods (columns Fret (in vivo) ```

Question 45

Descruve why grey scale camera are used for flouresecence microscopy

Answer 45

Colour cameras are not as sensitive

Question 46

What do dichroic mirrors relfect

Answer 46

Shorter wavelengths

Question 47

What is the light source used in flourescence microscopy

Answer 47

Laser or a mercury lamp

Question 48

What are the exictation and emission wavelengths of Blue flourescent protein ECFP

Answer 48

Exitate at 410 nm | emmit at 480 nm

Question 49

Descirbe the structure of a fluroescent fusion protein

Answer 49

Protein - peptide linker and flourophore

Question 50

Blue flouresecnet protein EXCITE COLOUR _______ EMITT COLOUR _______

Answer 50

violet Blue

Question 51

Green flouresecnet protein EXCITE COLOUR _______ EMITT COLOUR _______

Answer 51

Blue Green

Question 52

In Fret if there is no interaction what will be seen why will this happen

Answer 52

Violet light excites the BFP (blue) causes emission of blue light No GFP present so all observed is the blue light

Question 53

In FRET if there is an interaction describe what will be seen, why does this happen

Answer 53

Violet light excites the BFP - emission of blue light which then excites the GFP causing emssion of green light Green light is detect

Question 54

What does FRET stand for

Answer 54

Forster resonance energy transfer

Question 55

Describe how FRET may be used for protein and substrate

Answer 55

BFP - protein GFP - substrate If interaction between protein and sunstarate green light will be seen

Question 56

Describe how FRET may be used for a ligand

Answer 56

BFP and GFP either side of the ligand binding site If ligand binds these domains will be brought closer together If ligand binds - FRET

Question 57

Descibe how FRET may be used for a sensory domain

Answer 57

BFP - sensory doamin - substrate - GFP If substrate binds to sensory domain then BFP and GFP brought closer together and FRET

Question 58

Describe how FRET may be used with a proteease

Answer 58

GFP and BFP connected with a peptide linker If cleavage occurs there will be no FRET FRET will only occur if there is a link between the BFP and the GFP

Question 59

What type of afifnity do HSPGs have for RTKs

Answer 59

Low affinity