PKA Flashcards
What amino acids can be phosphorylated
serine, threonine, and tyrosine
What are the five basic steps of protein activity regulation?
- Ligand binding
- Second messanger
- Protein kinase cascade
- Transcription factor phosphorylation
- mRNA production
What is Protein Kinase A (PKA) activated by?
cAMP
True or False: All kinases bind to ATP and this is highly conserved?
True
True or False: cAMP is synthesized on demand?
True: cAMP levels are low until a. cyclase converts ATP into cAMP.
What does Adenylate Cyclase turn on?
it turns on the conversion from ATP to cAMP
What turns off cAMP production?
Phosphodiesterases
How is interaction specificity achieved?
A. cyclase controls flux of cAMP. this causes a localized rise in cAMP.
What are cAMP dependent protein kinases?
they are a family of PKAs that are activated by cAMP. They phosphorylate ser/thr hydroxyl group on target protein
What is the phosphorylation consensous for cAMP dependent Protein Kinases?
Arg-Arg-X-Ser
What Amino Acid can be used as a peptide inhibitor?
Alanine can be used as it is similar to Serine. It was used in crystal structure observations
What is highly conserved about PKA
*The proteins sequences/motifs.
*the Asp is highly conserved and involved in binding the terminal phosphate of ATP
What are some features of the crystal structure of PKA?
*the highly conserved ATP binding region
*they are bilobal with a hinge region that allow the unit to change from open/active to closed/inactive
*it has three conserved glycine residues on the N lobe that form a loop where ATP binds
*the terminal gamma phosphate of ATP interacts with a conserved Asp residue on the lower domain
What are the 4 parts of the PKA Catalytic subunit (C unit)
*N-lobe (has 3 gly residues)
* C-lobe (has substrate binding groove)
*Hinge region (opens and closes, allows ATP to bind)
*Catalytic cleft (where ATP is hydrolysis occurs)
*Substrate binding region
What are the physical properties of cAMP dependent protein kinases(CDPK)
its tetrameric protein with two regulatory subunits (R) and two catalytic subunits (C). When the protein is active the C subunits separate and the Two R units stay a dimer. The R units are a dimer even when inactive and are bound by a disulfide bond.
What does CDPK do?
it mediates the effects of the second messenger, cAMP.
What is the Mechanism of action for CDPK?
pseudosubstrate/ inhibitor domain
How is the PKA activated by cAMP?
The C subunits are bound to the R subunits via a substrate binding domain on the C unit that interacts with the pseudo-substrate domain on the R unit. 2 cAMP molecules (per R unit) co operatively bind to the R subunit, this weakens the interaction between the R and C units, causing them to dissociate from eachother. The now free C subunits can phosphorylate protein in their vacinity.
Once activated, what does a PKA do?
it often Phosphorylates TF in the nucleus. This is done by binding to the nucleus. it then activates CREB which turns on TF that lead to gene expression
What is an AKAP
An AKAP is an anchoring protein that contains multiple domains that interact with proteins and structures like the ER, this is its main function. It tethers proteins in place and allows for a localized production of cAMP. Its important in cross talk and bringing things together to create signaling complexes. it also acts as scaffolding.
What have structural studies taught us about how catalytic domains and their functions?
it taught us that the ATP binding region is conerved. that they are bilobal with a hinge region that allows for movement. that they contain 3 conserved glycine residues from form the ATP binding region. Look at hand drawn pic of the C and R subunits!
How is PKA activity regulated in space?
PKA is anchored to AKAPS. AKAPS tether the PKA to sub cellular structures like the ER and have a tail region that allows for binding to the R subunits of PKA. AKAPS bind other proteins associated with signal transduction such as PKC and phosphatases
