Cell Signaling Quiz 3 Flashcards
Why use binary switches?
Unambiguous
Straight forward reaction
Involve phosphorylation to drive the switch by protein kinases and protein phosphatases
When happens when an enzyme is turned on/off?
Change in enzymatic activity
Can be by conformation change
Could alter the primary structure and secondary structure
Could alter quaternary structure by dissociating polypeptides which make up protein
OR
Non- intrinsic modifications
ex. change in pH (like in the endomembrane system)
Phosphorylation
Explain reversibility to make the relationship regulated between glycogen and glucose?
Glycogen becomes glucose with phosphorylase
Glucose becomes glycogen with synthase
Cannot run at same time/need to be regulated
Therefore:
Phosphorylase is P’ed and off by default
Synthase is on by default and needs to be P’ed to turn it off
Kinase therefore inhibits synthase
Three classes of kinases
Ser/Thr - LOTS
Tyr - Some/Fewer
Dual-specificity - very few
Phosphatases of the three classes of kinases
Ser/Thr - few
Tyr - Plenty
Dual - Specificity - Some
Are kinases entergetically favorable?
Yes! Need to be for all of them!
ATP –> ADP + Pi
Reverse of this would waste too much energy and therefore would need to be performed by respiration
Phosphotase?
Hydrolysis rxn., break the phosphoester bond between A.A. and P.
Hydrolysis is thermodynamically favorable for tyrP and thermodynamically neutral for SerP and TheP
If hydrolysis is thermodynamically neutral for SerP and TheP, then why does the reaction happen?
Depends on the [reactants] & [products]
Molarity of H2O is 55.346 mol/L
Since there is normally plenty of water present, the forward reaction is favored by the high M of H2O
Conserved catalytic domain
Lysine in N-lobe
Aspartic acid in C-lobe
Both are used to maintain ATP in-between
Mutations in these A.A’s will have dead kinase
C-lobe of conserved catalytic domain
Aspartic acid
Contains activation loop which is regulatory and undergoes conformational change to activate kinase
Regulates hydrogen bonding of A.A. in active site, maintaining or disrupting active site
P’ion of active loop is common for kinase regulation
N-lobe of conserved catalytic domain
Lysine
Has C-helix which is there to control location of lysine. Ensures it interacts properly
This is critical to kinase activity
Nine families of kinases
Types of kinases:
1. Tyr (100)
2. Ser/Thr (400)
Therefore about 500 kinases in human genome
Families:
1. Tyr
2. AGC (PKA)
3. CAMK (Calmodulin dependent protein kinases)
4. CK1 (Casein kinases)
5. CMGC (CdK’s)
6. ST2 (MAP Kinases)
7. TKL (Raf)
8. RGC (receptor guanyl cyclase)
9. other (Includes polo kinases)
Ser/Thr (28)
PPP, PP1 (cacineurin, Fe3+, Zn2+, Mn+,) Use it as their ion
PPM, PP2C (Use Mg2+ as ion)
FCP, FCP1 (Use Mg2+ as ion, transcriptional regulaltion)
Tyr when cysteine is the key catalytic enzyme?
Class I:
PTP (receptor tyrosine phosphatases)
Dual Specificity (VH1, ~60)
Class II:
low molecular weight, PTP
LMPTP
Class III:
Cell cycle regulators (Cdc 25)
Class IV:
Haloacid dehydrogenases (EYA family) (eyes absent)
Tyr when Aspartic acid is the key catalytic enzyme?
Uses aspartic acid as nucleophile. Works in similar way as cysteine ones
Cysteine based Tyr
Similar catalytic domain
Have common motif (Cx5R)
C –> nucleophile will attack Tyr and form temporary phosphocysteine bond –> Water breaks this bond (hydrolyses it)
RTK Structure
N and C terminus
N to lipid bilayer is the extracellular receptor domain (highly variable)
Transmembrane domain
25-38 A.A.
Hydrophobic
Less variable
Under TMD have juxamembrane domain (JM)
Under JMD have the tyrosine kinase catalytic domain (TKC)
Under TKCD we have C-terminal tail area
RTK
Broad family with 58 genes and ~20 classes
Classic receptors of RTK classes?
Class I: EGFR
Class II: Insulin-R
Class III: PDGF: Platelet derived growth factor
Class IV: VEGF: Vascular endothelial growth factor (secreted to repair blood vessels)
Class V: FGF
Other receptors of RTK’s
Class IX: Ephr-R
Class XX: Undetermined
Class I RTK: ErbB or EGFR
4 members: ErB1, ErB2
Are associated with tumors and tumor progression
N-terminus: leucine-rich, cysteine-rich.
LR-1, CR-1, LR-2, CR-2
Class IV RTK: vEGF-R
N terminus: 7 Ig-like domains
Tyrosine kinase domain is split into 2
Ig-like domains:
vEGF-R1 which modulates vEGF-R2 activity levels
vEGF-R2 does all the work
Class V RTK: FGF
4 FGF genes have many versions
48 isoforms from splicing
N terminus: 3 Ig domains
D1-D2-D3
Different FGF molecules interact with different FGF receptors at different affinities
Only exception is FGF-7 which only interacts with FGFR2b
FGFR5 gene has no TK domain and could be a decoy
Class IX RTK: Ephrin-R
Cell-cell contact receptors
2 classes: EphrinA, EphrinB
EphrinA is a GPI anchor protein and is attached covalently to the PM
Therefore an enzyme can cut it and let GPI go into space
EphrinB is a transmembrane protein
Types of Eph receptors
EphR that binds EphA
EphR that binds EphB
There are 16 EphR (14 in humans)
EphRA 1-8, EphRA10
EphRB 1-4, EphRB6
Why does EphA have better bindng than EphB?
EphA/EphRA
EphB/EphRB
EphB has conformational change and makes process more complicated
What is Eph involved in?
Bidirectional signaling
Segmentation in development
Involved in somite formation:
-axonal migration via neurons
-key for cell migration
-angiogenesis formation of blood vessels
TH activation
Dimmerize –> cross
TH are towards each other
Activation loop
TK can cross P in many places
JM/TK/C-tail
P docking sites
Ways to specify protein bound
- Flanking with P like so
XXXYXXX
P - Maybe target for P. Maybe recruits other proteins
TK associated receptors
Most are cytokine receptors
Type I: N-Term has 4 helices. WSXWS.
W is tryptophan, S is serine, X is anything
These motifs are close 2 membranes
Type II: NO WSXWS
Type III: Immunoglobulin-family
JAK Janus Kinase
Jak1, Jak2, Jak3, Tyk2
120-140KDa
7 domains (JH)
N
JH4-7 (Ferm domain)
JH3 (SH2)
JH2 (pseudo kinase domain. No kinase activity. Maybe used to be something but is redundant now. If you remove it JH1 does not work. Maybe is structural. Maybe regulates JH1 and is conformational?)
JH1 (TK. Has JAK1-3 and TYK2)
What do JAKs recruit?
STATS which activates transcription proteins
STATs
7 STATs
STAT1-5, STAT5b, STAT6
80-90KDa
N
N-term (Short unclear structural function)
Coiled coil (alpha helix and alpha helix is stable rod. Contains NLS which is a nuclear localization sequence. STAT shuttles between nucleus and cytoplasm. Needs NLS to get in)
DNA binding domain
Linker domain (flexible)
SH2 (tyrosine binding domain. Can bind to P’ed tyrosine’s on JAK)
Transactivation domain (TAD, transactivation)
C
JAK-STAT pathway
Affinity for STAT-STAT P Y/SH2 binding is greater than affinity for STAT-JAK P Y/SH2 binding
Therefore STAT will dissociate from JAK after it is activated and will go to the nucleus, interact with other proteins and cause transcription
