2 Flashcards
What are receptors made of?
proteins
Since receptors are proteins, do ligands have to be proteins as well?
no
When is a protein said to be a target protein
when it’s activation results in a change in a cell- function, status, gene expression or shape
How many signalling proteins interact with target proteins?
one or more
What does a reaction cascade lead to?
the cascade leading to activation/ inhibition of target protein can either come as modification or interaction with the protein
once target protein is acted upon, this often leads to a feedback reaction that ges to the beginning of the signal
the receptors are often inhibited either by internalization or a change in the shape of the receptor
thus a signal not only acts on a target receptor but on a receptor itself
What are the 2 main classes of signal receptor pathways? What do they depend on?
- Pathways with cell-surface receptors- hydrophilic signal molecule + cell surface receptor
- Pathways with intracellular receptors- hydrophobic signal molecule + intracellular receptor
How many domains dose a cell surface receptor have? WHat are they and their functions?
3 domains
- extracellular domain which interacts with the signal
- cytoplasmic domain is the one that initiate the change inside the cell
- the portion within the bilipid bilayer is the transmembrane domain
Describe structure of transmembrane domain
- as the lipid biayer is lipid, the transmembrane portion is made of hydrophobic AA
each pass through the membrane is usually 20-24 AA in length- has several passes
hydrophobic AA that form transmembrane domain are called a-helixes
Describe structure of extracellular domain
- extracellular domain typically has disulphide bonds and CHO attachments that give it 3d structure to which hormones can come and interact
- extracellular domain is made of nitrogenous end which contains more cysteine which harbours disulphide binds
- disulphide bonds confer 3d structure and are also often glycosylated
Which domain contains disulphide bonds?
extracellular
Describe structure of cytoplasmic domain
made of COOH end of the protein
Are domains of receptors dependent on each other?
domains have modular nature- different domain form different receptors can be mis-matched-> domains can function independently from each other
Which protein end does signal bind to
Signal binds to ectodomain (NH2 end)
What are the potential problems of ectodomain-only receptors?
many receptors have isoforms that are purely made of extracellular domain which are directly released to the outside the cells
sometimes act as buffer systems for hormones, sometimes they don’t
e.g. GH has an isoform that has just the extracellular domain and acts as a buffer which increases the half-life of GH
sometimes this domain can stimulate antibody production which can end up at cell surface receptor and activate it.
This transmembrane receptor becomes active, leading to a continuous signal transduction
e.g. antibodies free extracellular domain for TSH receptor
releases TSH receptor stimulates antibody production which binds to THS receptor and stimulates thyroid gland-> hyperthyroidism-> Graves disease with increased thyroid hormone concentration-> goiter
What is the 1st step of the cascade?
the cytoplasmic domain, activated by conformational changes, induces a signalling cascade
what are conformational changes of the cytoplasmic domain induced by
- Phosphorylation of proteins
- Binding between proteins
Where and how does phosphorylation occur?
common post-translation modification is phosphorylation which typically occurs on the following AA: serine, threonine and tyrosine
Steps:
hydroxyl group is removed, phosphate group is added
phosphate group comes from ATP
What does phosphorylation of a protein result in?
in the conformational change of the protein
Does phosphorylation result in activation or deactivation?
Can lead to both
What is the function of phosphatases?
reverse phosphorylation (dephosphorylate)
Which AA out of 3 that get phosphorylated are more common?
Phosphorylated serines and threonines more abundant than phosphorylated tyrosines
Phosphorylation of which AA is usually the beginning of phosphorylation cascade?
Tyrosines phosphorylation often occurs at the beginning of a cascade (many receptors have or induce tyrosine kinase activity)
What do phosphorylated tyrosines usually act as?
The phosphorylated tyrosines serve as docking sites for down stream signal proteins- locking other protein at a phosphorylation site, keeping it activated
What are SH2 and SH3 domains?
The amino acid sequence that mediates docking to phosphorylated tyrosines (SH2 and SH3 domains) is conserved and diagnostic for proteins involved in the signaling cascade
SH2 domains allow proteins containing those domains to dock to phosphorylated tyrosine residues on other proteins. SH2 domains are commonly found in adaptor proteins that aid in the signal transduction of receptor tyrosine kinase pathways.
The classical SH3 domain is usually found in proteins that interact with other proteins and mediate assembly of specific protein complexes
What are the 3 major types of cell surface receptors?
1) Tyrosine kinase linked
i) Intrinsik TK activity- receptors themselves have enzymatic capability to phosphorylate tyrosine
ii) rectuited TK activity- recruit tyrosine kinase for phosphorylation to occur
2) G-protein coupled receptors
Which part of protein moves away after ligand binding? Why does it move away
G- alpha
Usually moves away to go on and activate an enzyme
Describe different types of G-alpha subunits
- Gs⍺ alpha: Activates adenylate cyclase- > uses cyclic AMP to activate PKA-> cascade reaction
- Gi⍺ alpha: Inhibits adenylate cyclase
- Gq⍺ alpha: Activates phospholipase C-> IP3 and DAG and Ca signaling
- Go⍺ alpha: Activates ion channels
- G12/13⍺: regulate Actin cytoskeleton-> G12/12a brings about a change in cell shape/movement of the cell
What is the most common type of cell surface receptros
g-protein
How many alpha-helixes form a pore of a g-protein receptors?
7 alpha- helix loops
Do endocrine signals or non-endocrine signals use G-protein receptors?
Both
- Many receptors of non-endocrine signals also act via G-proteins. Examples: glutamate (neurotransmitter), thrombin, odorants and photoreceptors
- Examples of endocrine hormone receptors are TRH, GnRH, TSH, LH, FSH, ACTH, GHRH and oxytocin
Do hormones only use one type of G-proteins
can use more than one
Is the type of g-protein used by the signalling molecule always constant?
G protein use may change during development
G protein use may change depending on the hormone concentration or in different tissues
Example of intrinsic TK receptor
insulin
Describe the structure of intrinsic TK receptors
has 2 domains- alpha and beta
alpha and beta are expressed by the same gene that has 3 portions: a, b , c
during post translation modification, c is cleaved off; a and b are joined by S-S bonds
b is transmembrane domain part
Which post-translation process does insulin receptor undergo?
Formation of disulfide bonds and proteolytic cleavage
Where is the highest concentration of insulin receptors found? Why?
highest in adipocytes and hepatocytes as these tissues are involved in glucose processing the most
Sequence of events after insulin binding
1) insulin binding to the receptor
2) Autophosphorylation of intracellular domain of receptor via intrinsic TK activity
3) Docking (recruitment) and phosphorylation of IRS-1 and IRS-2 (insulin receptor substrate)- dimerization
4) Activation of two major signal pathways- either MAPK or PI3 pathway
Insulin is similar to which hormone?
IGF-1
Insulin Receptor Signalling through MAPK pathway
1) insulin binds to it’s receptor and undergoes autophosphorylation on its carboxyl-terminal Tyr residues
2) Insulin receptor phosphorylates IRS-1
3) Grb2 binds to IRS-1 and SOS binds to GRB2, causing GDP release and GTP binding to Ras
4) Activated raf binds and activates Raf-1
5) Raf-1 phosphorylates MEK on 2 Ser residues, activating it. MEK phosphorylates ERK
6) ERK moves into the nucleus and phosphorylates nuclear transcription factors such as Elk1, activating them
7) Phosphorylated Elk1 joins SRF to stimulate the transcription and translation of a set of genes needed for cell division
What are Raf and Ras and when do they get activated?
Ras and Raf are enzymes which can act as kinases
activated when GDP is replaced with GTP
What are Grb2 and SOS
adaptor proteins
What are MEC and ERK
MEC and ERK are serine-threonine phosphorylases
spread the signal as they have multiple target- cytoplasmic protein or nuclear proteins
Which residues are phosphorylated in insulin receptor MAPK pathway
tyrosine residues, until Raf-1
Raf-1 phosphorylates Ser residues
MEK and ERK phosphorylate serine and threonine
Which protein is responsible for spreading the signal and entering the nucleus on MAPK pathway?
Phosphorylated Elk1
nsulin Receptor Signalling through PIP3
1) Irs-1 phosphorylates by insulin receptor activates PI3 kinase
2) PI3K phosphorylates a lipid by converting IP2 to IP3
3) PKB binds to PIP3 and begins PIP3 pathway
How does GSK3 act
Phosphorylated GsK3 is inactive
Not phosphorylated active GSK3 inactivates glycogen synthase (GS) by phosphorylating it
Active GS and NOT phosphorylated results in glycogen synthesis
How does PKB helps insulin depended glycogen synthesis process
brings glucose transporters to the cell surface
What are receptors with recruited tyrosine kinase activity characterized by
Characterized by 4 alpha-helices and homology of the ectodomain (hormone binding site)
What are receptors with recruited tyrosine kinase activity known for?
Best known are the receptors for growth hormone (GH), prolactin (PRL) and leptin
Receptors with recruited tyrosine kinase activity are a group of __/__ receptors
Receptors with recruited tyrosine kinase activity are a group of cytokine/haemopoietic receptors
Describe GH signalling pathway
1) GH has two binding sites and binds sequentially to two receptor molecules-> thus requires 2 receptors per molecule-> dimerization
2) Binds to 1 receptor first, which brings in 2nd receptor-> Dimerization of the cytoplasmic regions initiates signal transduction, by recruiting JAK-2 kinase
WHat initiate GH signalling pathway cascade?
JAK2 recruitment initiates phosphorylation cascade
What does JAK2 phosphorylate?
phosphorylates itself, the receptor and other proteins
What are the 3 branched of GH signalling pathways
- Branch 1: Activation of the transcription regulatory proteins, STAT, which are transcription factors (4 isoforms), found in the cytoplasm and move to the nucleus when activated
- Branch 2: Activation of the MAPK pathway (as insulin, but JAK2 plays the role of IRS1)
- Branch 3: Activation of PI3K pathway
Jak-stat pathway
1) after JAK is recruited and dimerization occurred,
tyrosine kinase activity is stimulated
2) they phosphorylate themselves and phosphorylate the receptors
3) this phosphorylation of the receptor’s cytoplasmic domain allows for recruitment of STAT (has 4 isoforms)
4) Stat is brought to the receptor
5) JAK-2 phosphorylates STAT
6) Phosphorylated STAT dimerize and are released from the receptor and go into the nucleus
7) Bind to DNA and effect gene expression as STATs are TF
Which pathways can G-protein can participate in?
Adenylyl cyclase
Phospholipase C
Which pathways can initiate MAPK or IP3 cascades?
IRS-1 or Jak-2
What is the length and the AA components of transmembrane domainÉ
20-25 a.a. needed to
cross membrane once (Ala, Ile, Leu, Met, Phe, Val, Pro, Gly)
